2-Aminopurine (2 AP) is a fluorescent isomer of adenine and has a fluorescence lifetime of ~11 ns in water. It is widely used in biochemical settings as a site-specific fluorescent probe of DNA and RNA structure and base-flipping and -folding. These assays assume that 2 AP is intrinsically strongly fluorescent. Here, we show this not to be the case, observing that gas-phase, jet-cooled 2-aminopurine and 9-methyl-2-aminopurine have very short fluorescence lifetimes (156 ps and 210 ps, respectively); they are, to all intents and purposes, non-fluorescent. We find that the lifetime of 2-aminopurine increases dramatically when it is part of a hydrate cluster, 2 AP · (H2O)n, where n = 1-3. Not only does it depend on the presence of water molecules, it also depends on the specific hydrogen-bonding site to which they attach and on the number of H2O molecules at that site. We selectively microhydrate 2-aminopurine at its sugar-edge, cis-amino or trans-amino sites and see that its fluorescence lifetime increases by 4, 50 and 95 times (to 14.5 ns), respectively.

Intersystem crossing rates of S 1 state keto-amino cytosine at low excess energy The amino-keto tautomer of supersonic jet-cooled cytosine undergoes intersystem crossing (ISC) from the v = 0 and low-lying vibronic levels of its S 1 ( 1 ππ * ) state. We investigate these ISC rates experimentally and theoretically as a function of S 1 state vibrational excess energy E exc . The S 1 vibronic levels are pumped with a ∼5 ns UV laser, the S 1 and triplet state ion signals are separated by prompt or delayed ionization with a second UV laser pulse. After correcting the raw ISC yields for the relative S 1 and T 1 ionization cross sections, we obtain energy dependent ISC quantum yields Q corr ISC = 1%-5%. These are combined with previously measured vibronic state-specific decay rates, giving ISC rates k ISC = 0.4-1.5 · 10 9 s −1 , the corresponding S 1 S 0 internal conversion (IC) rates are 30-100 times larger. Theoretical ISC rates are computed using SCS-CC2 methods, which predict rapid ISC from the S 1 ; v = 0 state with k ISC = 3 · 10 9 s −1 to the T 1 ( 3 ππ * ) triplet state. The surprisingly high rate of this El Sayed-forbidden transition is caused by a substantial admixture of 1 n O π * character into the S 1 ( 1 ππ * ) wave function at its non-planar minimum geometry. The combination of experiment and theory implies that (1) below E exc = 550 cm −1 in the S 1 state, S 1 S 0 internal conversion dominates the nonradiative decay with k IC ≥ 2 · 10 10 s −1 , (2) the calculated S 1 T 1 ( 1 ππ * 3 ππ * ) ISC rate is in good agreement with experiment, (3) being El-Sayed forbidden, the S 1 T 1 ISC is moderately fast (k ISC = 3 · 10 9 s −1 ), and not ultrafast, as claimed by other calculations, and (4) at E exc ∼ 550 cm −1 the IC rate increases by ∼50 times, probably by accessing the lowest conical intersection (the C5-twist CI) and thereby effectively switching off the ISC decay channels. C 2015 AIP Publishing LLC. [http://dx

Ultraviolet radiative damage to DNA is inefficient because of the ultrafast S1 ⇝ S0 internal conversion of its nucleobases. Using picosecond pump-ionization delay measurements, we find that the S1((1)ππ*) state vibrationless lifetime of gas-phase keto-amino cytosine (Cyt) is τ = 730 ps or ∼ 700 times longer than that measured by femtosecond pump-probe ionization at higher vibrational excess energy, Eexc. N1-Alkylation increases the S1 lifetime up to τ = 1030 ps for N1-ethyl-Cyt but decreases it to 100 ps for N1-isopropyl-Cyt. Increasing the vibrational energy to Eexc = 300-550 cm(-1) decreases the lifetimes to 20-30 ps. The nonradiative dynamics of S1 cytosine is not solely a property of the amino-pyrimidinone chromophore but is strongly influenced by the N1-substituent. Correlated excited-state calculations predict that the gap between the S2((1)nOπ*) and S1((1)ππ*) states decreases along the series of N1-derivatives, thereby influencing the S1 state lifetime.

We have investigated the S0 → S1 UV vibronic spectrum and time-resolved S1 state dynamics of jet-cooled amino-keto 1-methylcytosine (1MCyt) using two-color resonant two-photon ionization, UV/UV holeburning and depletion spectroscopies, as well as nanosecond and picosecond time-resolved pump/delayed ionization measurements. The experimental study is complemented with spin-component-scaled second-order coupled-cluster and multistate complete active space second order perturbation ab initio calculations. Above the weak electronic origin of 1MCyt at 31 852 cm−1 about 20 intense vibronic bands are observed. These are interpreted as methyl group torsional transitions coupled to out-of-plane ring vibrations, in agreement with the methyl group rotation and out-of-plane distortions upon 1ππ∗ excitation predicted by the calculations. The methyl torsion and ν1′ (butterfly) vibrations are strongly coupled, in the S1 state. The S0 → S1 vibronic spectrum breaks off at a vibrational excess energy Eexc ∼ 500 cm−1, indicating that a barrier in front of the ethylene-type S1⇝S0 conical intersection is exceeded, which is calculated to lie at Eexc = 366 cm−1. The S1⇝S0 internal conversion rate constant increases from kIC = 2 ⋅ 109 s−1 near the S1(v = 0) level to 1 ⋅ 1011 s−1 at Eexc = 516 cm−1. The 1ππ∗ state of 1MCyt also relaxes into the lower-lying triplet T1 (3ππ∗) state by intersystem crossing (ISC); the calculated spin-orbit coupling (SOC) value is 2.4 cm−1. The ISC rate constant is 10–100 times lower than kIC; it increases from kISC = 2 ⋅ 108 s−1 near S1(v = 0) to kISC = 2 ⋅ 109 s−1 at Eexc = 516 cm−1. The T1 state energy is determined from the onset of the time-delayed photoionization efficiency curve as 25 600 ± 500 cm−1. The T2 (3nπ∗) state lies >1500 cm−1 above S1(v = 0), so S1⇝T2 ISC cannot occur, despite the large SOC parameter of 10.6 cm−1. An upper limit to the adiabatic ionization energy of 1MCyt is determined as 8.41 ± 0.02 eV. Compared to cytosine, methyl substitution at N1 lowers the adiabatic ionization energy by ≥0.32 eV and leads to a much higher density of vibronic bands in the S0 → S1 spectrum. The effect of methylation on the radiationless decay to S0 and ISC to T1 is small, as shown by the similar break-off of the spectrum and the similar computed mechanisms.

The decay mechanism of S 0 → S 1 excited cytosine (Cyt) and the effect of substitution are studied combining jet-cooled spectroscopy (nanosecond resonant two-photon ionization (R2PI) and picosecond lifetime measurements) with CASPT2//CASSCF computations for eight derivatives. For Cyt and five derivatives substituted at N1, C5, and C6, rapid internal conversion sets in at 250−1200 cm −1 above the 0 0 0 bands. The break-off in the spectra correlates with the calculated barriers toward the "C5−C6 twist" conical intersection, which unambiguously establishes the decay mechanism at low S 1 state vibrational energies. The barriers increase with substituents that stabilize the charge shifts at C4, C5, and C6 following ( 1 ππ*) excitation. The R2PI spectra of the clamped derivatives 5,6-trimethyleneCyt (TMCyt) and 1-methyl-TMCyt (1M-TMCyt), which decay along an N3 out-of-plane coordinate, extend up to +3500 and +4500 cm −1 .