Radiation damage in DNA: possible role of higher triplet states.

Triplet states of deoxyribose are expected to dissociate efficiently into radicals, leading to strand breaks. Such states could be excited by slow secondary electrons (A) or result from ion recombination in spurs containing two or more ion pairs (B). Estimates of the efficiencies of these processes are presented and the mechanisms discussed in the light of recent work with electrons, vacuum ultraviolet (VUV) photons, and X rays. Route B could play a significant role in producing double-strand breaks, while route A may be a better approach to characterizing the process experimentally.

The effects of weak magnetic fields on radical recombination reactions in micelles.

PURPOSE: To demonstrate the effects of weak magnetic fields (> approximately 1 mT) on chemical reactions involving free radicals, in the context of possible effects of environmental electromagnetic radiation on biological systems. MATERIALS AND METHODS: Transient absorption, flash photolysis experiments have been performed to study the kinetics and yields of radical reactions. The triplet state of benzophenone has been used as a convenient source of radical pairs, whose identity is largely immaterial to the investigation of the so-called Low Field Effect. Hydrogen abstraction from surfactant molecules in micelles yields a pair of neutral radicals, one large and one small, in a region of restricted translational and rotational motion. RESULTS: In alkyl sulphate and sulphonate micelles a weak field increases the concentration of free radicals that escape from the micelle to an extent that depends on the structure, dynamics and volume of the space in which the radical pairs are confined. The effect (up to 10%) is typically largest at 1-2 mrT. Smaller effects are found for Brij and TX100 micelles. CONCLUSIONS: Low Field Effects depend strongly on the local environment of the radical pair. Larger effects than observed here might be expected for radicals formed from singlet (rather than triplet) precursors, as would be the case in biological reactions.

Critical energies for SSB and DSB induction in plasmid DNA by low-energy photons: action spectra for strand-break induction in plasmid DNA irradiated in vacuum.

PURPOSE: To measure action spectra for the induction of single-strand breaks (SSB) and double-strand breaks (DSB) in plasmid DNA by low-energy photons and provide estimates for the energy dependence of strand-break formation important for track-structure simulations of DNA damage. MATERIALS AND METHODS: Plasmid pMSG-CAT was irradiated as a monolayer, under vacuum, with 7 150eV photons produced by a synchrotron source. Yields of SSB and DSB were determined by the separation of the three plasmid forms by gel electrophoresis. RESULTS: The yields of SSB per incident photon increased from 1.4x 10(-15) SSB per plasmid per photon/cm2 at 7eV to 7.5 x 10(-14) SSB per plasmid per photon/cm2 at 150 eV. Direct induction of DSB was also detected increasing from 3.4 x 10(-17) DSB per plasmid per photon/cm2 at 7eV to 4.1 x 10(-15) DSB per plasmid per photon/cm2 at 150eV. When the absorption cross-section of the DNA was considered, the quantum efficiency for break formation increased over the energy range studied. Over the entire energy range, the ratio of SSB to DSB remained constant. CONCLUSIONS: These studies provide evidence for the ability of photons as low as 7 eV to induce both SSB and DSB. The common action spectrum for both lesions suggests that they derive from the same initial photoproducts under conditions where the DNA is irradiated in vacuum and a predominantly direct effect is being observed. The spectral and dose-effect behaviour indicates that DSB are induced predominantly by single-event processes in the energy range covered.

Critical energies for ssb and dsb induction in plasmid DNA by vacuum-UV photons: an arrangement for irradiating dry or hydrated DNA with monochromatic photons.

PURPOSE: Theoretical modelling techniques are often used to simulate the action of ionizing radiations on cells at the nanometre level. Using monoenergetic vacuum-UV (VUV) radiation to irradiate DNA either dry or humidified, the action spectra for the induction of DNA damage by low energy photons and the role of water and can be studied. These data provide inputs for the theoretical models. METHODS: Various combinations of monochromator, grating and VUV window have been used to obtain monochromatic photons from the 2 GeV electron synchrotron at the CLRC, Daresbury Laboratory. A sample chamber containing plasmid DNA is installed at the end of the beamline. The chamber can be evacuated or water can be introduced (as water vapour or humidified helium). In this way, DNA can be irradiated either dry or humidified. RESULTS: An arrangement for irradiating dry or humidified DNA using monoenergetic photons from 7 eV to 150 eV has been developed. At the energies used, exposure rates vary from about 5 x 10(10) to 3 x 10(12) photons cm(-2) s(-1) over a 1 cm2 sample area. At all but the lowest energies this is sufficient to produce significant levels of DNA damage in just a few minutes. The measured dose variation over the sample area is typically 30%, but this is reduced significantly using sample scanning techniques.

Magnetic isotope effects in biology: a marker for radical pair reactions and electromagnetic field effects?

PURPOSE: To explore the possible usefulness of magnetic isotope effects, especially to detect processes affected by magnetic fields. MATERIALS AND METHODS: The theoretical model of Brocklehurst and McLauchlan is used to estimate the effects of isotopic substitution (e.g. 13C for 12C) on the yields of chemical reactions involving radical pairs. RESULTS: It is demonstrated that in some circumstances isotope effects on the yields from initially singlet pairs (bond breaking to give radicals) could be very large and easily detected. The effects on radical pairs formed by random encounters are necessarily much smaller but may be detectable. Illustrative calculations are presented for carbon, hydrogen and oxygen isotopes; deuterium substitution also produces very large mass effects but its study could be useful to support work on carbon. CONCLUSIONS: Effects may be small but worth looking for: distinguishing them from mass effects will be aided by measuring distributions within product molecules. In laboratory experiments (especially tomography) a combination of isotopic substitution and applied fields is a potentially powerful technique.

Free radical mechanism for the effects of environmental electromagnetic fields on biological systems.

The radical pair mechanism is discussed as a possible route whereby a magnetic field of environmental strength might affect a biological system. It is well established as the origin of reproducible field effects in chemistry, and these can be observed even at very low magnetic field strengths, including that of the geomagnetic field. Here it is attempted to give a description which might assist experimentalists working in biological laboratories to devize tests of its relevance to their work. The mechanism is well understood and a specific theoretical approach is taken to explore and emphasize the importance of the lifetime of the radical pair and the precise chemical natures of the radicals which comprise it in affecting the size of the low-field effects. Further subsequent processes are likely necessary to cause this primary effect to attain biological significance. Arguments are provided to suggest that the encounters of freely diffusing pairs (F-pairs) of radicals are unlikely to produce significant effects in biology.

Action spectra for single- and double-strand break induction in plasmid DNA: studies using synchrotron radiation.

Ionizing radiations deposit a wide range of energies in and around DNA and this leads to a corresponding spectrum of complexity of the lesions induced. The relationships between the amount of energy deposited and the yields and types of damage induced are important in modelling the physical and chemical stages of radiation effect and linking them to biological outcome. To study these relationships experimentally, plasmids were mounted as a monolayer and exposed in vacuum to near-monoenergetic photons from the Daresbury Synchrotron. After irradiation, the DNA was washed off and assayed for single-(ssb) and double-strand breaks (dsb) using agarose gel electrophoresis. Dose-effect relationships for ssb and dsb induction were obtained at various energies in the range 8-25 eV. The initial responses in the low-dose region allowed damage yields to be estimated. However, a common feature is that the responses showed energy-dependent plateaus at higher doses as if a fraction of the DNA were shielded. Various measures were taken both to minimize and to correct for this effect. The data appear to show that the yields of ssb and dsb increase only slowly with photon energies > 10 eV, with a suggestion of similar threshold energies for both lesions. In the energy range covered, the yield of ssb is 12-20-fold greater than that of dsb. The data indicate that ssb and dsb may have a common precursor in this system. Earlier work with low-energy electrons showed that at 25 eV ssb were induced but no dsb were detected.