Quantitative analysis of UV photolesions suggests that cyclobutane pyrimidine dimers produced in mouse skin by UVB are more mutagenic than those produced by UVC.

The amount of photolesions produced in DNA after exposure to physiological doses of ultraviolet radiation (UVR) can be estimated with high sensitivity and at low cost through an immunological assay, ELISA, which, however, provides only a relative estimate that cannot be used for comparisons between different photolesions such as cyclobutane pyrimidine dimer (CPD) and pyrimidine(6-4)pyrimidone photoproduct (64PP) or for analysis of the genotoxicity of photolesions on a molecular basis. To solve this drawback of ELISA, we introduced a set of UVR-exposed, calibration DNA whose photolesion amounts were predetermined and estimated the absolute molecular amounts of CPDs and 64PPs produced in mouse skin exposed to UVC and UVB. We confirmed previously reported observations that UVC induced more photolesions in the skin than UVB at the same dose, and that both types of UVR produced more CPDs than 64PPs. The UVR protection abilities of the cornified and epidermal layers for the lower tissues were also evaluated quantitatively. We noticed that the values of absorbance obtained in ELISA were not always proportional to the molecular amounts of the lesion, especially for CPD, cautioning against the direct use of ELISA absorbance data for estimation of the photolesion amounts. We further estimated the mutagenicity of a CPD produced by UVC and UVB in the epidermis and dermis using the mutation data from our previous studies with mouse skin and found that CPDs produced in the epidermis by UVB were more than two-fold mutagenic than those by UVC, which suggests that the properties of CPDs produced by UVC and UVB might be different. The difference may originate from the wavelength-dependent methyl CpG preference of CPD formation. In addition, the mutagenicity of CPDs in the dermis was lower than that in the epidermis irrespective of the UVR source, suggesting a higher efficiency in the dermis to reduce the genotoxicity of CPDs produced within it. We also estimated the minimum amount of photolesions required to induce the mutation induction suppression (MIS) response in the epidermis to be around 15 64PPs or 100 CPDs per million bases in DNA as the mean estimate from UVC and UVB-induced MIS.

In†Situ Raman Monitoring of Silver(I)-Aided Laser-Driven Cleavage Reaction of Cyclobutane.

The cyclobutane cleavage reaction is an important process and has received continuous interest. Herein, we demonstrate the visible laser-driven cleavage reaction of cyclobutane in crystal form by using in situ Raman spectroscopy. Silver(I) coordination-induced strain and thermal effects from the laser irradiation are the two main driving forces for the cleavage of cyclobutane crystals. This work may open up a new avenue for studying cyclobutane cleavage reactions, as compared to the conventional routes using ex situ techniques.

An electro-optical and electron injection study of benzothiazole-based squaraine dyes as efficient dye-sensitized solar cell materials: a first principles study.

Squaraine dyes have attracted significant attention in many areas of daily life from biomedical imaging to semiconducting materials. Moreover, these dyes are used as photoactive materials in the field of solar cells. In the present study, we investigated the structural, electronic, photophysical, and charge transport properties of six benzothiazole-based squaraine dyes (Cis-SQ1-Cis-SQ3 and Trans-SQ1-Trans-SQ3). The effect of electron donating (-OCH3) and electron withdrawing (-COOH) groups was investigated intensively. Ground state geometry and frequency calculations were performed by applying density functional theory (DFT) at B3LYP/6-31G** level of theory. Absorption spectra were computed in chloroform at the time-dependent DFT/B3LYP/6-31G** level of theory. The driving force of electron injection (ΔG (inject)), relative driving force of electron injection (ΔG r (inject)), electronic coupling constants (|VRP|) and light harvesting efficiency (LHE) of all six compounds were calculated and compared with previously studied sensitizers. The ΔG (inject), ΔG r (inject) and |VRP| of all six compounds revealed that these sensitizers would be efficient dye-sensitized solar cell materials. Cis/Trans-SQ3 exhibited superior LHE as compared to other derivatives. The Cis/Trans geometric effect was studied and discussed with regard to electro-optical and charge transport properties.

Ring opening of the cyclobutane in a thymine dimer radical anion.

The reactions of hydrated electrons (e(aq) (-)) with thymine dimer 2 and thymidine have been investigated by radiolytic methods coupled with product studies, and addressed computationally by means of BB1K-HMDFT calculations. Pulse radiolysis revealed that one-electron reduction of the thymine dimer 2 affords the radical anion of thymidine (5) with t(1/2)<35 ns. Indeed, the theoretical study suggests that radical anion 3, in which the spin density and charge distribution are located in both thymine rings, undergoes a fast partially ionic splitting of the cyclobutane with a half-life of a few ps. This model fits with the in vivo observation of thymine dimer repair in DNA by photolyase. gamma-Radiolysis of thymine dimer 2 demonstrates that the one-electron reduction and the subsequent cleavage of the cyclobutane ring does not proceed by means of a radical chain mechanism, that is, in this model reaction the T(-)* is unable to transfer an electron to the thymine dimer 2.

Detailed dynamics of the photodissociation of cyclobutane.

Semiclassical electron-radiation-ion dynamics simulations are reported for the photodissociation of cyclobutane into two molecules of ethylene. The results clearly show the formation of the tetramethylene intermediate diradical, with dissociation completed in approximately 400 fs. In addition, the potential energy surfaces of the electronic ground state and lowest excited-state were calculated at the complete-active-space self-consistent-field/multireference second-order perturbation theory (CASSCF/MRPT2) level with 6-31G* basis sets, along the reaction path determined by the dynamics simulations. There are well-defined energy minima and maxima in the intermediate state region. It is found that both C-C-C bond bending and rotation of the molecule (around the central C-C bond) have important roles in determining the features of the potential energy surfaces for the intermediate species. Finally, the simulations and potential energy surface calculations are applied together in a discussion of the full mechanism for cyclobutane photodissociation.

Amino substituted bisketenes: generation, structure, and reactivity.

Hitherto unknown diamino-substituted bisketenes with both free (14) and tethered (16) amino substituents have been generated by using laser flash photolysis for ring opening of the corresponding cyclobutenediones. The time-resolved kinetics of ring closure of the amino bisketenes back to the cyclobutenediones were measured by IR or UV spectroscopy, and give first-order rate constants which vary by a factor of 7.5x10(4), and the bis(Me2N) bisketene 14 is the most reactive in ring closure that has been reported. Rate constants for ring closure of these and previously observed bisketenes vary by a factor of 10(13). The dialkylamino bisketenes 16 (R=Me, n-Bu) with tethered substituents and restricted geometries are less reactive than the bis(Me2N) bisketene 14 by factors of 1700 and 540, respectively. Computational results obtained with DFT methods suggest angle strain in the tethered cyclobutenediones 15 inhibits facile cyclization of bisketenes 16.

Photodissociation of cyclobutyl bromide at 234 nm studied using velocity map imaging.

This study investigates the 234 nm photodissociation dynamics of cyclobutyl bromide using a two-dimensional photofragment velocity imaging technique. The spin-orbit ground- and excited-state Br(2P) atoms are state-selectively detected via [2+1] resonance enhanced multiphoton ionization (REMPI), whereas the cyclobutyl radicals are ionized using 157 nm laser light. The Br(2P(3/2)) and the Br(2P(1/2)) atoms and their c-C4H7 radical cofragments evidence a single-peaked, Gaussian-shaped translational energy distribution ranging from approximately 14 to approximately 39 kcal/mol and angular distributions with significant parallel character. The Br(2P(1/2))/ Br(2P(3/2)) spin-orbit branching ratio is determined to be 0.11 +/- 0.07 by momentum match between the Br(2P) photofragments and the recoiling c-C4H7 fragments, assuming a uniform photoionization probability of the c-C4H7 radicals with an internal energy range of 10-35 kcal/mol. The REMPI line strength ratio for the detection of Br(2P(3/2)) and Br(2P(1/2)) atoms at 233.681 and 234.021 nm, respectively, is therefore derived to be 0.10 +/- 0.07. The measured recoil kinetic energies of the c-C4H7 radicals, and the resulting distribution of internal energies, indicates some of the radicals are formed with total internal energies above the barrier to isomerization and subsequent dissociation, but our analysis indicates they may be stable due to the substantial fraction of the internal energy which is partitioned to rotational energy of the radicals.

Experimental determination of the heat of hydrogenation of phenylcyclobutadiene.

The heat of hydrogenation of phenylcyclobutadiene (DeltaH degrees (hyd) = 57.4 +/- 4.9 kcal mol(-1)) was determined via a thermodynamic cycle by carrying out gas-phase measurements on 1-phenylcyclobuten-3-yl cation. This leads to an antiaromatic destabilization energy of 27 +/- 5 kcal mol(-1), a difference of 9.6 +/- 4.9 kcal mol(-1) for the first and second C-H bond dissociation energies of 1-phenylcyclobutene, and an estimate of 96 +/- 5 kcal mol(-1) for the heat of formation of cyclobutadiene. These results are compared to G3, G3(MP2), and B3LYP computations and represent the first experimental measurements of the energy of a monocyclic cyclobutadiene.

Bipyrimidine photoproducts rather than oxidative lesions are the main type of DNA damage involved in the genotoxic effect of solar UVA radiation.

Exposure to solar UV radiation gives rise to mutations that may lead to skin cancer. UVA (320-340 nm) constitutes the large majority of solar UV radiation but is less effective than UVB (290-320 nm) at damaging DNA. Although UVA has been implicated in photocarcinogenesis, its contribution to sunlight mutagenesis has not been elucidated, and DNA damage produced by UVA remains poorly characterized. We employed HPLC-MS/MS and alkaline agarose gel electrophoresis in conjunction with the use of specific DNA repair proteins to determine the distribution of the various classes and types of DNA lesions, including bipyrimidine photoproducts, in Chinese hamster ovary cells exposed to pure UVA radiation, as well as UVB and simulated sunlight (lambda > 295 nm) for comparison. At UVA doses compatible with human exposure, oxidative DNA lesions are not the major type of damage induced by UVA. Indeed, single-strand breaks, oxidized pyrimidines, oxidized purines (essentially 8-oxo-7,8-dihydroguanine), and cyclobutane pyrimidine dimers (CPDs) are formed in a 1:1:3:10 ratio. In addition, we demonstrate that, in contrast to UVB and sunlight, UVA generates CPDs with a large predominance of TT CPDs, which strongly suggests that they are formed via a photosensitized triplet energy transfer. Moreover, UVA induces neither (6-4) photoproducts nor their Dewar isomers via direct absorption. We also show that UVA photons contained in sunlight, rather than UVB, are implicated in the photoisomerization of (6-4) photoproducts, a quickly repaired damage, into poorly repaired and highly mutagenic Dewar photoproducts. Altogether, our data shed new light on the deleterious effect of UVA.

On the roles of urocanic acid in photoimmunosuppression: attempted photorepair of urocanic acid-DNA cyclobutane adducts with DNA photolyase.

It has been reported that UV-induced immunosuppression can be reversed by photoreactivation or exposure to T4 endonuclease V, two treatments that can repair cyclobutane pyrimidine dimers. These observations, together with the known role of urocanic acid (UA) in UV-induced immune suppression, prompted us to study the ability of DNA photolyase to repair UA-DNA cyclobutane photoadducts in single-stranded calf thymus DNA. We did not detect any release of UA, with a sensitivity implying that photolyase is at least 2900 times less active toward UA-DNA adducts than toward cis-syn thymine-thymine dimers. This indicates that any reversal of photoimmunosuppression by photoreactivation cannot significantly involve cleavage of UA-DNA cyclobutane adducts.

Repair of cyclobutane pyrimidine dimers and 6-4 photoproducts in the fission yeast Schizosaccharomyces pombe.

We have measured repair of both of the major lesions induced by ultraviolet irradiation (cyclobutane pyrimidine dimers and 6-4 photoproducts) in wild-type Schizosaccharomyces pombe and in selected rad mutants, including mutants with deletions in genes from the main phenotypic groups. We find that rad13 delta, rad15 and rad16 delta, which are the S. pombe homologues of the excision-defective Saccharomyces cerevisiae rad2, rad3 and rad1, respectively, repair lesions somewhat more slowly than the wild type, but still have considerable repair capacity. rad2 delta, also a presumed excision-defective mutant, behaves similarly. rad8 and rad9 delta, which belong to different phenotypic groups, repair lesions at the same rate as wild-type cells. These findings provide new evidence that S. pombe has a second repair system for removing ultraviolet damage, which is absent in S. cerevisiae. Surprisingly, this second mechanism repairs lesions very efficiently; its possible nature is discussed.

Relative induction of cyclobutane dimers and cytosine photohydrates in DNA irradiated in vitro and in vivo with ultraviolet-C and ultraviolet-B light.

SV40 DNA was irradiated in vitro and in vivo with UV-C (240-280 nm) and UV-B (280-320 nm) light, and damaged sites sensitive to digestion with Escherichia coli endonuclease III (endo III) and bacteriophage T4 endonuclease V (endo V) were quantified. The frequency of endo III-sensitive sites (primarily cytosine photohydrates) induced was 1-2% of the frequency of endo V-sensitive sites (cyclobutane dimers) in both purified SV40 DNA and intracellular episomal SV40 DNA. Endo III- and endo V-sensitive sites in DNA were induced in the same relative proportion at both UV-C and UV-B wavelengths. We found no evidence to support earlier inferences that intracellular conditions enhance the formation of cytosine photohydrates or other monobasic forms of DNA damage.

The base-alteration spectrum of spontaneous and ultraviolet radiation-induced forward mutations in the URA3 locus of Saccharomyces cerevisiae.

A forward mutation system has been developed to obtain rapidly clonable mutants at the URA3 locus in yeast by means of selection for 5-fluoroorotic acid resistance. We have used this system to determine base changes in 35 spontaneous and 34 ultraviolet radiation-induced ura3 base substitution mutants. Other mutants (frameshift, deletion, duplication, replacement) were detected as well. Evidence is reported which suggests cyclobutane dimers are the principal mutagenic lesions induced by UV radiation in stationary phase cells of the yeast Saccharomyces cerevisiae. Since most of the induced lesions are at 5'-TT-3' sites, the results suggest that the "A-rule", preferential insertion of adenine residues opposite poorly pairing sites in DNA, does not apply for yeast cells irradiated in stationary phase, whereas the spontaneous mutation data indicate that the A-rule applies for cells in logarithmic phase. Most of the spontaneous mutations are transversions. UV-induced transitions and transversions occur at approximately equal frequencies.

Cyclobutane-type pyrimidine photodimer formation and excision in human skin fibroblasts after irradiation with 313-nm ultraviolet light.

The formation and excision of 313-nm light-induced cyclobutane-type pyrimidine photodimers were determined in confluent cultures of human fibroblasts. A new method was developed for the resolution and determination of cytosine-thymine (CT) and thymine-thymine dimers (TT) by using sodium borohydride reduction and high-pressure liquid chromatography. This assay can detect as little as 1.8 TT or 5.6 CT per 10(8) daltons, levels induced in monolayers of human skin fibroblasts by doses of 1 and 2 kJ m-2 of 313-nm light, respectively. CT formation was 20% more efficient than TT formation in the physiological dose range of 2.25-15 k m-2 at 37 degrees C. Normal fibroblasts removed 61% TT within the first 8 h of incubation following a dose of 5.5 kJ m-2. CT was removed approximately twice as efficiently as TT during the same time period following exposure to 10 kJ m-2. The lack of removal of CT as well as TT observed in xeroderma pigmentosum fibroblasts indicates that the repair deficiency in these cells affects the repair of both classes of dimers.