Remarkable effects of solvent and substitution on the photo-dynamics of cytosine: a femtosecond broadband time-resolved fluorescence and transient absorption study.

Cytosine (Cyt) among all the nucleic acid bases features the most complex and least understood nonradiative deactivation, a process that is crucially important for its photostability. Herein, the excited state dynamics of Cyt and a series of its N1- and C5-derivatives, including the full set of Cyt nucleosides and nucleotides in DNA and RNA and the nucleosides of 5-methyl cytosine, 5-methylcytidine and 2'-deoxy-5-methylcytidine, have been investigated in water and in methanol employing femtosecond broadband time-resolved fluorescence coupled with fs transient absorption spectroscopy. The results reveal remarkable state-specific effects of the substitution and solvent in tuning distinctively the timescales and pathways of the nonradiative decays. For Cyt and the N1-derivatives, the nonradiative deactivations occur in a common two-state process through three channels, two from the light-absorbing ππ* state with respectively the sub-picosecond (∼0.2 ps) and the picosecond (∼1.5 ps) time constant, and the third is due to an optically dark nπ* state with the lifetime ranging from several to hundreds of picoseconds depending on solvents and substitutions. Compared to Cyt, the presence of the ribose or deoxyribose moiety at the N1 position of N1-derivatives facilitates the formation of the nπ* at the sub-picosecond timescale and at the same time increases its lifetime by ∼4-6 times in both water and methanol. In sharp contrast, the existence of the methyl group at the C5 position of the C5-derivatives eliminates completely the sub-picosecond ππ* channel and the channel due to the nπ*, but on the other hand slows down the decay of the ππ* state which after relaxation exhibits a single time constant of ∼4.1 to ∼7.6 ps depending on solvents. Varying the solvent from water to methanol accelerates only slightly the decay of the ππ* state in all the compounds; while for Cyt and its N1-derivatives, this change of solvent also retards strongly the nπ* channel, prolongs its lifetime from such as ∼7.7 ps in water to ∼52 ps in methanol for Cyt and from ∼30 ps in water to ∼186 ps in methanol for deoxycytidine. The spectral signatures we obtained for the ππ* and the nπ* states allow unambiguous evidence for clarifying uncertainties in the excited states of Cyt and the derivatives. The results provide a unifying experimental characterization at an improved level of detail about the photophysics of Cyt and its analogues under biologically relevant conditions and may help in understanding the photostability as well as photo-damages of the bases and related DNAs.

The solvent effect and identification of a weakly emissive state in nonradiative dynamics of guanine nucleosides and nucleotides--a combined femtosecond broadband time-resolved fluorescence and transient absorption study.

A combined method of femtosecond broadband time-resolved fluorescence (fs-TRF) and transient absorption (fs-TA) was employed to investigate the excited state dynamics of 2'-deoxyguanosine (dG) and 2'-deoxyguanosine 5'-monophosphate (dGMP). Comparative fs-TRF and fs-TA measurements were conducted on dG and dGMP in neutral water, deuterated water, and methanol with excitation wavelengths of 245, 267 and 285 nm. Very similar results were observed with dG and dGMP. The data provide compelling evidence for the co-existence of two nonradiative pathways. One is the generally recognized Laππ* mediated channel, the other involves an unprecedented weakly emissive state which plays a significant role in the overall deactivation processes. The Laππ* channel features biphasic dynamics with time constants (τ1/τ2) of ~0.2/0.8 ps in water and ~0.25/1.0 ps in methanol. The biphasic decay arises due to a partial transfer with τ1 of the Laππ* population to the newly identified state followed by conversion in τ2 of the remaining Laππ* molecules into the electronic ground state. The channel mediated by the weakly emissive species shows solvent-dependent dynamics with time constants (τ3) of ~2.0 ps in water, ~2.3 ps in deuterated water, and ~4.1 ps in methanol. The species features absorption at UV wavelengths (~300-400 nm) and exhibits deeply red-shifted fluorescence (λmax ~ 520 nm) with polarization direction varied markedly from that of the Laππ* but close to the Lbππ*. This species acts as an effective quenching state to the radiative decay of the brightly emissive Laππ* and Lbππ*. It sets in promptly (<~50 fs) after the photoexcitation and is further populated through nonadiabatic coupling with the Laππ*. The overall involvement of this state is facilitated with excitation at high energy and is favoured in methanol over water. According to the spectral character and the solvent effect in particular the kinetic isotope effect, the species is tentatively associated to the πσ* state with charge transfer (CT) character which is considered to be preferentially stabilized by hydrogen-bonding between the guanine amino and surrounding solvent molecules. The result of this study leads to a dramatically different picture of guanine deactivation. It demonstrates a crucial role of the solvent in shaping the nonradiative dynamics of guanine nucleosides and nucleotides. The data presented are important for understanding the detailed photophysics of not only the monomeric guanine but also DNA assemblies that contain guanine in base pairs or have a guanine tetrad as the structural motif.

Femtosecond broadband time-resolved fluorescence and transient absorption study of the intramolecular charge transfer state of methyl 4-dimethylaminobenzoate.

A combined application of femtosecond broadband time-resolved fluorescence (fs-TRF), fluorescence anisotropy (fs-TRFA) and fs to microsecond (µs) transient absorption (TA) have been used to probe directly the dynamics, nature, formation and decay paths of the singlet intramolecular charge transfer ((1)ICT) state of methyl 4-dimethylaminobenzoate (1a) in acetonitrile. The result reveals explicit evidence for a common electronic origin (the L(a) nature) of the (1)ICT state and its precursor the locally excited ((1)LE) state to account jointly for the dual florescence known to this system. It also shows that the ICT reaction from the (1)LE to (1)ICT state occurs with time constant of ~0.8 ps and the (1)ICT state formed decays with a ~1.9 ns time constant leading mainly to a ππ* natured triplet state ((3)T(1)). The (3)T(1) then relaxes with a ~4 µs lifetime under deoxygenated condition resulting in full recovery of the ground state (S(0)). As a case study, this work contributes novel experimental data for improved understanding of the mechanism of ICT reaction; it also reveals a distinct deactivation pattern for this prototype para-amino substituted aromatic carbonyl compound in acetonitrile.

A triphenylphosphonium-functionalised cyclometalated platinum(II) complex as a nucleolus-specific two-photon molecular dye.

An organometallic cyclometalated platinum(II) complex, [Pt(L(3))Cl][PF(6)], has been synthesised from a specially designed cyclometalating ligand, HL(3) (triphenyl{5-[3-(6-phenylpyridin-2-yl)-1H-pyrazol-1-yl]pentyl}phosphonium chloride), that contains a pendant carbon chain carrying a terminal cationic triphenylphosphonium moiety. Aside from its room temperature single-photon luminescent properties in solution, [Pt(L(3))Cl](+) can also produce two-photon-induced luminescence at room temperature upon excitation at 700 nm from a mode-locked Ti:sapphire laser. Its two-photon absorption cross-section in DMF at room temperature was measured to be 28.0x10(-50) cm(4) s photon(-1). [Pt(L(3))Cl](+) is able to selectively stain the cell nucleolus. This has been demonstrated by two-photon confocal imaging of live and methanol-fixed HeLa (human cervical carcinoma) and 3T3 (mouse skin fibroblasts) cells. This organelle specificity is likely to be related to its special affinity for proteins within cell nucleoli. As a result of such protein affinity, [Pt(L(3))Cl](+) is an efficient RNA transcription inhibitor and shows rather profound cytotoxicity. On the other hand, the organelle-specific labelling and two-photon-induced luminescent properties of [Pt(L(3))Cl](+) renders it a useful nuclear dye for the 3-dimensional reconstruction of optical sections of thick tissues, for example, mouse ileum tissues, by multiphoton confocal microscopy.