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J. Phys. Chem. B 2007, 111, 5518-5524
Triplet Formation of 4-Thiothymidine and Its Photosensitization to Oxygen Studied by Time-Resolved Thermal Lensing Technique Yosuke Harada,† Tadashi Suzuki,*,† Teijiro Ichimura,† and Yao-Zhong Xu‡ Department of Chemistry and Materials Science, Tokyo Institute of Technology, 2-12-1 Ohokayama, Meguro-ku, Tokyo 152-8551, Japan, and Department of Chemistry, the Open UniVersity, Milton Keynes, MK7 6AA, United Kingdom ReceiVed: NoVember 23, 2006; In Final Form: March 9, 2007
Excited-state dynamics of 4-thiothymidine (S4-TdR) and its photosensitization to molecular oxygen in solution with UVA irradiation were investigated. Absorption and emission spectra measurements revealed that UVA photolysis of S4-TdR gives rise to a population of T1(ππ*), following S2(ππ*) f S1(nπ*) internal conversion. In transient absorption measurement, the 355 nm laser photolysis gave broad absorption (380-600 nm) bands of triplet S4-TdR. The time-resolved thermal lensing (TRTL) signal of S4-TdR containing the thermal component due to decay of triplet S4-TdR was clearly observed by the 355 nm laser excitation. The quantum yield for S1 f T1 intersystem crossing was estimated to be unity by a triplet quenching experiment with potassium iodide. In the presence of molecular oxygen, the photosensitization from triplet S4-TdR gave rise to singlet oxygen O2 (1∆g) with a quantum yield of 0.50. Therapeutic implications of such singlet oxygen formation are discussed.
Introduction DNA is the fundamental genetic material. Naturally occurring DNA, unlike protein, does not contain sulfur. Sulfur-containing thio-analogues of DNA building blocks such as thio-bases and thio-nucleosides have attracted considerable attention since the isolation of 4-thiouridylic acid from Escherichia coli by Lipsett.1 They possess various distinct properties relative to normal DNA constituents.2-6 In particular, the high sensitivity of thio-bases to UVA light enables one to target DNA containing the thioanalogues selectively while minimizing any effect on normal DNA, because normal bases (A, G, C, T), the most important chromophores of DNA, are transparent in the UVA region. Synergism between thio-analogues and UVA light is being explored to offer a new therapeutic approach for cancer and other disease.7-10 4-Thiothymidine (S4-TdR) is an analogue of the naturally occurring nucleoside thymidine (TdR) in which the oxygen atom at the 4-position is replaced by a sulfur atom (see Chart 1) and this thio-analogue has strong absorption in the UVA region.9,11 Massey et al. suggested a potential therapeutic use for S4-TdR combined with UVA light.12 They found that S4-TdR can be readily incorporated into cellular DNA, and that low, normally nontoxic doses of UVA can easily inflict lethal damage to S4TdR containing DNA. The nature of the lethal DNA lesion(s) and the mechanism by which S4-TdR and UVA cause cell death is still unknown. However, it is reasonable to expect that formation of an electronically excited state of S4-TdR is present at the initial and crucial stage of UVA-induced cell killing. Thus, photophysical and photochemical studies of S4-TdR are likely to provide insights into the mechanisms of action of S4-TdR in the UVA-induced cell killing. * To whom correspondence should be addressed. Phone/fax: +81-35734-2331. E-mail:
[email protected]. † Tokyo Institute of Technology. ‡ The Open University.
CHART 1: Molecular Structure of TdR (X ) O) and S4-TdR (X ) S)
We investigated excited-state dynamics of S4-TdR by means of absorption and emission spectroscopy, transient absorption measurement, and the time-resolved thermal lensing (TRTL) technique. In particular, we utilized the TRTL method to elucidate the notable photophysical properties of S4-TdR: the triplet formation process and the photosensitization to molecular oxygen. The TRTL method is a photothermal technique that provides a powerful tool with which to explore nonradiative deactivation processes of photoexcited molecules.13-17 In this report, we present the novel photoactive properties of S4-TdR and discuss its photomechanisms and therapeutic potential. Experimental Details Materials. 4-Thiothymidine (S4-TdR) was synthesized as previously reported.18 Thymidine (TdR) (Aldrich) was used as received. 2-Hydroxybenzophenone (2-HBP) (Kanto Chemical) was purified by repeated recrystalization from methanol and n-hexane. Acetonitrile and ethanol (Kanto Chemical, GR grade) were used as the solvent without further purification. For transient absorption and TRTL measurements, sample solutions were deaerated by bubbling with Ar gas (99.95%) prior to
10.1021/jp0678094 CCC: $37.00 © 2007 American Chemical Society Published on Web 04/18/2007
Triplet Formation of 4-Thiothymidine measurements, used without deaeration, or flushed with O2 gas for 10 min. The sample was then passed into a cell (optical path length 1 cm). Apparatus. The UV absorption spectrum was measured with a double beam spectrometer (Jasco Ubest V-550). Emission and excitation spectra were measured by a spectrofluorometer (Jasco FP-6500) equipped with a quartz Dewar at 77 K with liquid nitrogen. Transient absorption measurements were carried out with a Nd:YAG laser (Continuum Surelite II; 355 nm, 6 ns pulse duration,
