QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in aqueous solution

QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in aqueous solution | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in aqueous solution Binbin Xie, Zhao Rui, Jia-Ling Dai, Bo-Wen Yin, Li Shuai This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6547757/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Selenonucleobases have garnered increasing interest from both experimental and theoretical communities for their promising roles in photodynamic therapy and DNA crosslinking. Similar to the extensively investigated thymidine:4-thiothymidine system, the selenium-modified thymidine:4-selenothymidine (Tp 4 Se T) dimer may also exhibit significant photochemical activity within DNA duplexes. However, its detailed photochemical reaction mechanisms remain largely unexplored. Herein, we employed high-level MS-QM(CASPT2//CASSCF) method to explore excited-state decay, [2 + 2] cycloaddition and (6 − 4) reactions of Tp 4 Se T in aqueous solution. Our calculations revealed five possible nonadiabatic decay channels enabling population of the T 1 state from the initial S 2 state, mediated by two multi-state intersections of S 2 /S 1 /T 2 /T 1 and S 1 /T 2 /T 1 . Following population of the T 1 state, the [2 + 2] cycloaddition proceeds via a stepwise, nonadiabatic mechanism. That is, the pathway starts from Tp 4 Se T in the T 1 state via the T 1CC or T 1CSe intermediates and ultimately ends up with Se 5 -selenetane in the S 0 state. Subsequent transformation of Se 5 -selenetane into the Se 5 -(6 − 4) product occurs through a concerted reaction in the ground state, characterized by the simultaneous cleavage of the C 4 –Se 8 bond and formation of the S 8 –H 9 bond. This study provides detailed mechanistic insights into the photoreactivity of selenonucleobases in DNA duplexes at the molecular level. Physical sciences/Chemistry/Theoretical chemistry/Computational chemistry Physical sciences/Chemistry/Physical chemistry/Excited states Physical sciences/Chemistry/Photochemistry/Photobiology Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 1. Introduction Among various phototherapeutic approaches, photodynamic therapy has emerged as a clinically approved and effective modality that utilizes light to activate photosensitizers (PSs) — photoactive compounds capable of producing reactive oxygen species (ROS). 1 , 2 Upon photoexcitation to their triplet states, PSs mediate ROS production through two primary pathways: Type I mechanisms involve electron/hydrogen transfer with biomolecules to form superoxide anions (O 2 − ) or hydroxyl radicals (·OH), while Type II mechanisms utilize triplet-triplet energy transfer to convert ground-state oxygen ( 3 O 2 ) into cytotoxic singlet oxygen ( 1 O 2 ). These ROS species play pivotal roles in selective apoptosis induction for therapeutic applications spanning antimicrobial treatment, dermatological disorders, and oncological interventions. 3 , 4 Current clinically approved PSs predominantly comprise porphyrin derivatives and transition metal complexes, where metal incorporation enhances triplet quantum yields. However, challenges persist regarding oxygen dependency (particularly problematic in hypoxic tumor microenvironments) and non-specific phototoxicity toward healthy tissues. 5 , 6 Addressing these limitations necessitates rational design of novel PSs with optimized ROS generation efficiency, hypoxia tolerance, and biological selectivity — goals requiring fundamental understanding of photophysical processes (e.g., excited-state lifetimes, intersystem crossing efficiency) and photochemical reactions within biological contexts. To circumvent metal-induced cytotoxicity, recent efforts have focused on developing metal-free PS systems. 7 , 8 Sulfur-substituted PSs have emerged as promising candidates due to their tunable photophysics. 9 – 11 Strategic substitution of carbonyl oxygen with sulfur atoms in molecular frameworks enables enhanced intersystem crossing (ISC) and near-unity triplet quantum yields. In particular, thionucleobases have garnered significant attention as photosensitizers owing to their structural resemblance to natural nucleobases, along with several advantageous properties: (1) excellent biocompatibility, (2) red-shifted absorption profiles that facilitate deeper tissue penetration, (3) large photon absorption cross-sections, and (4) strong spin-orbit coupling combined with small singlet–triplet energy gaps, enabling highly efficient triplet-state population. 12 – 27 Expanding this strategy to heavier chalcogens, selenonucleobases have recently gained experimental and theoretical interest. 28 – 45 Crespo-Hernández and colleagues utilized time-resolved spectroscopy to probe the photochemical properties of 6-thioguanine and its selenium analogue, 6-selenoguanine. 38 Their comparative analysis revealed that replacing sulfur with selenium leads to a further red-shift in absorption and promotes ISC, though it also results in a relatively short triplet-state lifetime. From a theoretical perspective, Pirillo et al. conducted DFT and TD-DFT computations to evaluate the photophysical characteristics, including absorption profiles, excited-state energies, and spin–orbit coupling elements, of thymidine and deoxyguanosine substituted with selenium and tellurium. 39 , 46 Their findings indicate that these compounds exhibit efficient ISC and possess adequate triplet energies to activate molecular oxygen, suggesting their promise as UVA-targeted therapeutic agents. Additionally, Hazra and Manae investigated the influence of electronic conjugation and chalcogen atomic size on the spectral and reactive properties of modified thymine molecules. 47 In parallel, Mai and collaborators carried out multireference electronic structure calculations and nonadiabatic dynamics simulations at the MS-CASPT2 and ADC(2) levels, unveiling rapid population and depopulation processes of the reactive triplet state in 2-selenouracil. 36 More recently, Fang’s group applied MS-CASPT2 and combined QM(MS-CASPT2)/MM methods to examine the excited-state relaxation mechanisms of 6-selenoguanine in different environments, including gas phase, aqueous solution, and DNA matrices. 34 , 37 Besides, Peng et al. systematically explored the photophysical behavior of 2-selenouracil, 4-selenouracil, and 2,4-diselenouracil through MS-CASPT2 calculations, highlighting the position-dependent influence of selenium substitution on excited-state dynamics. 35 Furthermore, Valverde et al. used QM/MM dynamics to study 6-selenoguanine in water and found an ISC time of 452 fs, longer than the experimental 130 fs, which reflects multiple overlapping processes rather than ISC alone. 29 While these studies have offered valuable mechanistic insights into selenonucleobases, current researches have primarily focused on their electronic structures and nonadiabatic dynamics related to photophysical behavior. In contrast, their photochemical reactions remain insufficiently characterized. To fill this gap, we employed high-level complete active space self-consistent field (CASSCF) and its multistate second-order perturbation extension (MS-CASPT2) with in the quantum mechanics/molecular mechanics (QM/MM) framework to investigate the excited-state relaxation pathways, [2 + 2] photocycloaddition, and subsequent (6 − 4) photoproduct formation of the Tp 4 Se T dimer in aqueous solution (Scheme 1 ). In addition, we further compared these systems with thymine:4-thiothymine (Tp 4 S T) to assess selenium substitution effects on both photophysical and photochemical behaviors of Tp 4 Se T. 48 2. Computational details 2.1 System setup To account for solvation effects, a spherical water box (radius = 25 Å) containing 1977 water molecules was generated using Packmol. 49 The Tp 4 Se T dimer was placed at the center of this box. A quick energy minimization (2000 steps) was performed to eliminate unphysical molecular contacts, followed by a 50 ps MD simulation at 300 K using a Nóse–Hoover thermostat. During the simulation, water molecules beyond 20 Å from the Tp 4 Se T center of mass were frozen. The Tp 4 Se T dimer was modeled using the all-atom CHARMM general force field, 50 , 51 while the solvent molecules were described using the TIP3P model. 52 The final MD snapshot was used to prepare the QM/MM system by removing fixed waters, resulting in a 20 Å solvation shell with 3144 atoms (Fig. 1 ). All simulations were conducted using CHARMM (version c31b1). 53 2.2 QM/MM calculations Following equilibration MD simulations, all subsequent electronic structure calculations, including geometry optimizations and single-point energy refinements, were performed using the high-level complete active space self-consistent field (CASSCF) and its multistate second-order perturbation extension (MS-CASPT2) with in the QM/MM framework. 54 – 56 In this hybrid scheme, the Tp 4 Se T dimer (30 atoms) was treated quantum mechanically at MS-CASPT2//CASSCF level, while the MM subsystem employed the TIP3P water model. 52 The simulation system consisted of 2694 mobile atoms (within 15 Å of the Tp 4 Se T dimer) and 450 fixed peripheral atoms. The electronic structure calculations employed an active space comprising sixteen electrons distributed across twelve orbitals for both CASSCF and MS-CASPT2 treatments ( Figure S1 ). Key computational parameters included: (1) Cholesky decomposition with unbiased auxiliary basis sets for efficient two-electron integral evaluation; 57 (2) zero ionization potential-electron affinity (IPEA) shift; 58 and (3) an imaginary shift of 0.2 atomic units to mitigate intruder-state artifacts. 59 Spin-orbit coupling (SOC) constants were computed using the atomic mean-field approximation (AMFI) approach: 60 $$\:⟨{\psi\:}_{I}\left|{H}_{eff}^{SOC}\right|{\psi\:}_{J}⟩\:=\:\sqrt{\:[⟨{\psi\:}_{I}|{H}_{x}^{SOC}|{\psi\:}_{J}⟩²\:+\:⟨{\psi\:}_{I}|{H}_{y}^{SOC}|{\psi\:}_{J}⟩²\:+\:⟨{\psi\:}_{I}|{H}_{z}^{SOC}|{\psi\:}_{J}⟩²]\:/\:3}\:$$ All quantum mechanical calculations, including both QM(CASSCF)/MM and QM(MS-CASPT2)/MM approaches, were carried out using the aug-cc-pVDZ basis set for the tellurium atom and the cc-pVDZ basis set for all other atoms. 61 , 62 These calculations were implemented through the OpenMolcas program (version v21.02) 63 in conjunction with the TINKER molecular modeling package (version 6.3.2). 64 3. Results and Discussion 3.1 Excited-State Decay of Tp 4 Se T Vertical excitation properties. We initially performed geometry optimization for the ground-state minimum of Tp 4 Se T at the QM(CASSCF)/MM level, which is denoted as S 0 in Fig. 2 . On this optimized structure, vertical excitation energies and electronic configurations of the low-lying excited states, namely, S 1 ( 1 n H π L+1 *), S 2 ( 1 π H−1 π L+1 *), T 1 ( 3 π H−1 π L+1 *), T 2 ( 3 n H π L+1 *), and T 3 ( 3 π H−4 π L *), were examined based on QM(MS-CASPT2)/MM calculations (see Table 1 ). Frontier molecular orbital analysis indicates that these excitations are predominantly localized on the 4-selenothymine moiety. Specifically, the HOMO corresponds to a nonbonding n orbital centered on the Se atom (Se 8 ), the HOMO-1 is attributed to the π orbital of the C 4 = Se 8 bond, and the LUMO + 1 is a delocalized π* orbital spanning the 4-selenothymine ring ( Figure S1 ). The S 1 state arises primarily from a HOMO → LUMO + 1 transition, with a dominant configuration weight of 0.56. As expected, this state is optically forbidden with a negligible oscillator strength. The QM(MS-CASPT2)/MM method predicted vertical excitation energy of 58.5 kcal/mol for the S 1 state, which is 5.0 kcal/mol lower than the value reported by Pirillo et al. 39 In contrast, the S 2 state is optically allowed, involving a major HOMO-1 → LUMO + 1 transition (weight: 0.67), and exhibits a significantly higher oscillator strength. The computed vertical excitation energy for the S₂ state (85.3 kcal/mol) at the Franck–Condon geometry in a water environment is consistent with the value reported by Pirillo et al., who obtained 83.0 kcal/mol for 4-selenothymidine in water using the IEFPCM model. 39 Additionally, Shi et al. experimentally determined the maximum absorption band of 4-selenothymidine to be 77.7 kcal/mol using UV spectroscopy, which is slightly lower than the values predicted by theoretical calculations. 32 This discrepancy may be attributed to the fact that, as noted by Götze et al., vertical excitation energies are typically higher than experimental absorption maxima, since the latter reflect transitions from vibrationally relaxed states. 65 Additionally, the vertical excitation energies for the T 1 ( 3 π H−1 π L+1 *) and T 2 ( 3 n H π L+1 *) states were calculated to be 55.4 and 56.6 kcal/mol, respectively, which are in good agreement with the values reported by Pirillo et al. at the M06/6–31 + G* level. 46 The electronic configurations of T 1 and T 2 correspond to ππ* and nπ* characters, respectively. The T 3 ( 3 π H−4 π L *) state exhibits a significantly higher vertical excitation energy—approximately 5.0 kcal/mol above that of the S 2 state. This might indicate that the T 3 state plays a relatively minor role during the excited-state relaxation process. Table 1 QM(MS-CASPT2//CASSCF)/MM computed vertical excitation energies (kcal/mol) of the lowest several excited singlet and triplet states of Tp4SeT. States MS-CASPT2 M06/6–31 + G* a Exp. b S 1 ( 1 n H π L+1 *) 58.5 63.5 S 2 ( 1 π H−1 π L+1 *) 85.3 83.0 77.7 T 1 ( 3 π H−1 π L+1 *) 55.4 50.7 T 2 ( 3 n H π L+1 *) 56.6 58.6 T 3 ( 3 π H−4 π L *) 90.0 — a Theor. Chem. Acc. 2016, 135, 8; b Nucleos. Nucleot. Nucl. 2021, 40, 96–116. Adiabatic excitation properties. As illustrated in Fig. 2 , we optimized the minimum-energy geometries of Tp 4 Se T in the S 1 ( 1 nπ * ), S 2 ( 1 ππ * ), T 1 ( 3 ππ * ), T 2 ( 3 nπ * ), and T 3 ( 3 ππ * ) states at the QM(CASSCF)/MM level without imposing any geometric constraints. The electronic transitions and associated structural rearrangements are mainly localized on the 4-selenothymine chromophore, except for the T 3 minimum, where the excitation is primarily centered on the thymine unit. Figure 3 presents the bond length differences between these five excited-state minima and the ground-state minimum. The most significant structural changes are observed in the C 4 –Se 8 , C 4 –C 5 , and C 5 –C 6 bond lengths, while variations in other bonds remain relatively minor compared to the S 0 structure. For example, in the S 2 ( 1 ππ * ) minimum, the C 4 –Se 8 bond is markedly elongated to 1.989 Å, approximately 0.2 Å longer than in the S 0 minimum. Additionally, the C 4 –C 5 bond shortens by over 0.12 Å, while the C 5 –C 6 bond lengthens by more than 0.11 Å. The structural variations in the S 1 ( 1 nπ * ), T 1 ( 3 ππ * ), and T 2 ( 3 nπ * ) minima exhibit similar trends relative to S 0 , with bond length variations generally confined within the range of − 0.12 to 0.12 Å. The adiabatic excitation energies corresponding to the T 1 ( 3 ππ * ), T 2 ( 3 nπ * ), T 3 ( 3 ππ * ), S 1 ( 1 nπ * ), and S 2 ( 1 ππ * ) minima were computed to be 46.1, 49.6, 76.4, 48.4, and 66.2 kcal/mol, respectively, at the QM(MS-CASPT2)/MM level (Table 2 ). Notably, at the S 1 ( 1 nπ * ) minimum geometry, the single-point energies of the S 1 ( 1 nπ * ), T 1 ( 3 ππ * ), and T 2 ( 3 nπ * ) states are nearly degenerate, with values of 48.4, 48.1, and 47.6 kcal/mol, respectively. This near-degeneracy implies that the S 1 ( 1 nπ * ) minimum could serve as a key intersection point in the excited-state relaxation pathways. In contrast, the relatively high adiabatic excitation energy associated with the T 3 ( 3 ππ * ) minimum suggests that this state is less likely to contribute significantly to the excited-state decay dynamics. Table 2 QM(MS-CASPT2//CASSCF)/MM calculated relative energies (in kcal/mol) of minima of Tp4SeT. The potential energy of the S0 minimum is taken as the reference zero point. Structures Energies Structures Energies S 0 0.0 T 1 46.1 S 1 48.4 T 2 49.6 S 2 66.2 T 3 76.4 Several surface-crossing structures involving the S 0 , S 1 ( 1 nπ * ), S 2 ( 1 ππ * ), T 1 ( 3 ππ * ), and T 2 ( 3 nπ * ) states of Tp 4 Se T were identified through QM(CASSCF)/MM calculations. These include three minimum-energy conical intersections between singlet (S 2 /S 1 and S 1 /S 0 ) and triplet (T 2 /T 1 ) states, along with four singlet–triplet crossing points (S 0 /T 1 , S 1 /T 1 , S 1 /T 2 , and S 2 /T 2 ). The optimized geometries and corresponding bond length variations for these conical intersections and crossing points are depicted in Figs. 3 and 4 . Nearly all intersection structures exhibit substantial geometric distortions compared to the S 0 minimum. In particular, pronounced bond length changes, ranging from − 0.16 to 0.35 Å, are observed for key bonds such as C 4 –Se 8 , N 3 –C 4 , C 4 –C 5 , and C 5 –C 6 . Interestingly, only the S 1 /T 1 and S 1 /T 2 , intersection structures retain an essentially planar configuration. By contrast, the T 2 /T 1 and S 1 /S 0 conical intersections show slight out-of-plane displacement of the Se 8 atom relative to the pyrimidine ring. Moreover, prominent pyramidalization of carbon or nitrogen atoms is observed in the S 2 /S 1 , S 0 /T 1 , and S 2 /T 2 crossings, indicating strong out-of-plane distortions associated with the surface-crossing geometries. Energetically, the S 2 /S 1 and S 1 /S 0 conical intersections are located at 83.6/80.9 and 70.4/66.2 kcal/mol, respectively. Given that the vertical excitation energy of S 2 is 85.3 kcal/mol, both intersections are readily accessible, thus promoting efficient internal conversion pathways back to the ground state. Notably, at the S 2 /S 1 conical intersection, the S 1 , S 2 , T 1 and T 2 states are nearly degenerate, forming a unique four-state crossing. This feature may facilitate both internal conversion and intersystem crossing. The T 2 /T 1 conical intersection lies at a relatively low energy (51.8/50.2 kcal/mol), suggesting that rapid internal conversion from T 2 to T 1 to is likely to occur through this intersection. Regarding the crossing points, the single-point energies of S 2 /T 2 are calculated to be 89.7/88.6 kcal/mol, approximately 4.0 kcal/mol higher than the vertical excitation energy at the S 2 Franck–Condon point. This indicates that access to the S 2 /T 2 crossing is energetically unfavorable, and thus it likely plays a minor role in the excited-state deactivation processes. In contrast, the S 1 /T 2 and S 1 /T 1 crossing structures can be regarded as three-state intersections (S 1 /T 2 /T 1 and S 1 /T 1 /T 2 ), with single-point energies of 57.5/53.9/53.2 and 54.8/54.2/54.2 kcal/mol, respectively. As expected, the S 0 /T 1 crossing structure exhibits a relatively higher energy of 67.1/68.3 kcal/mol. This observation aligns with the experimentally reported high triplet yield in the overall excited-state relaxation dynamics. Table 3 QM(MS-CASPT2//CASSCF)/MM calculated relative energies (in kcal/mol) of conical intersections and crossing points of Tp4SeT. The potential energy of the S0 minimum is taken as the reference zero point. Structures Energies Structures Energies S 1 /S 0 70.4/66.2 S 1 /T 1 b 54.8/54.2 S 2 /S 1 a 83.6/80.9 S 1 /T 2 b 57.5/53.9 T 2 /T 1 51.8/50.2 S 2 /T 2 89.7/88.6 S 0 /T 1 67.1/68.3 a S₂/S₁ forms a four-state crossing (S 1 , S 2 , T 1 and T 2 ) with close energies: 83.6, 80.9, 81.6, and 79.3 kcal/mol; b S₁/T₁ and S₁/T₂ are three-state crossings (S 1 , T 1 and T 2 ) with energies of 54.8/54.2/54.2 and 57.5/53.9/53.2 kcal/mol. Excited-state decay pathways. To gain further insight into the efficient triplet-state population in Tp 4 Se T, we performed linearly interpolated internal coordinate (LIIC) path calculations at the QM(MS-CASPT2)/MM level of theory (Figs. 5 and S2). Starting from the S 2 Franck–Condon region, five potential nonadiabatic decay pathways were identified along the LIIC profiles: Path I: S 2 ( 1 ππ * )-FC → S 2 /S 1 → S 1 → S 1 /T 1 /T 2 → T 1 ;Path II: S 2 ( 1 ππ * )-FC → S 2 /S 1 → S 1 → S 1 /T 2 /T 1 → T 2 → T 2 /T 1 → T 1 ; Path III: S 2 ( 1 ππ * )-FC → S 2 /S 1 /T 2 /T 1 → S 1 → S 1 /S 0 → S 0 . Path IV: S 2 ( 1 ππ * )-FC → S 2 /S 1 /T 2 /T 1 → T 1 ; Path V: S 2 ( 1 ππ * )-FC → S 2 /S 1 /T 2 /T 1 → T 2 → T 2 /T 1 → T 1 . Table 4 QM(MS-CASPT2)/MM calculated spin-orbit couplings (in cm− 1) at several important intersection structures of Tp4SeT. Structures States SOCs Structures States SOCs SOCs a S 2 /S 1 S 2 -T 2 306.6 S 0 /T 1 S 0 -T 1 41.6 — S 2 -T 1 160.6 S 1 /T 1 S 1 -T 1 425.0 891.5 S 1 -T 2 162.9 S 1 /T 2 S 1 -T 2 17.8 17.3 S 1 -T 1 394.3 S 2 /T 2 S 2 -T 2 346.0 303.7 a Theor. Chem. Acc. 2016, 135, 8. We begin by discussing the excited-state decay pathway that involves only internal conversion, namely Path III. As shown in the left panel of Fig. 5 , this pathway requires overcoming an energy barrier of approximately 14.0 kcal/mol from the S 2 minimum to the S 2 /S 1 conical intersection. The relative energy at this conical intersection (83.6/80.9 kcal/mol) remains below that of the S 2 Franck–Condon region (85.3 kcal/mol), indicating that the initially populated S 2 ( 1 ππ * ) state can undergo internal conversion to the S 1 ( 1 nπ * ) state near this region due to the small energy gap. Subsequently, the S 1 ( 1 nπ * ) population can decay to the ground state via the S 1 /S 0 conical intersection, although this process requires overcoming an energy barrier of approximately 20 kcal/mol. The right panel of Fig. 5 illustrates a similar initial process, where the population transitions from the initially excited S 2 ( 1 ππ * ) state to the S 1 ( 1 nπ * ) state via the S 2 /S 1 conical intersection. Unlike Path I, the decay pathways of Paths II and III are more likely to proceed via intersystem crossing to the T 2 ( 3 nπ * ) or T 1 ( 3 ππ * ) states through the S 1 /T 1 /T 2 or S 1 /T 2 /T 1 crossings. This is facilitated by their near-degeneracy and the significant SOCs of 425.0 cm − 1 (S 1 -T 1 ) and 17.8 cm − 1 (S 1 -T 2 ). Subsequently, the T 2 ( 3 nπ * ) state can further relax to the lower-lying T 1 ( 3 ππ * ) state via the T 2 /T 1 conical intersection. As illustrated in Figure S2 , transitions from S 2 ( 1 ππ * ) to the reactive T 1 ( 3 ππ * ) state via the four-state intersection of S 2 /S 1 /T 2 /T 1 are also accessible (Paths IV and V). These processes benefit from both efficient internal conversion and intersystem crossing. At this multistate intersection, the molecule may directly transition to T 1 ( 3 ππ * ) (Path IV), aided by a S 2 -T 1 SOC of 160.6 cm − 1 , or proceed indirectly via intermediate T 2 ( 3 nπ * ) state with a S 2 -T 2 SOC of 306.6 cm − 1 (Path V). From this intermediate, further relaxation to T 1 ( 3 ππ * ) is highly probable due to T 2 → T 1 internal conversion. The proposed excited-state decay pathways above are consistent with El-Sayed's rule, 66 which states that ISC is significantly enhanced when it involves a change in orbital character. For instance, the SOC constants between S 2 ( 1 ππ * ) and T 2 ( 3 nπ * ) at the S 2 /T 2 crossing, and between S 1 ( 1 nπ * ) and T 1 ( 3 ππ * ) at the S 1 /T 1 crossing were estimated to be 346.0 and 425.0 cm − 1 , respectively. Notably, at the S 2 /S 1 /T 2 /T 1 multi-state intersection, the electronic configurations of both singlet (S 2 , S 1 ) and triplet (T 2 , T 1 ) states exhibit strong mixing of nπ * and ππ * characters. This leads to exceptionally large SOCs not only for S 2 -T 2 and S 1 -T 1 , but also for S 2 -T 1 and S 1 -T 2 transitions. Taking into account both the relative energetics and the computed SOC constants, we concluded that the ultrafast and energetically accessible nonadiabatic decay pathways efficiently promote triplet-state population in Tp 4 Se T. In contrast, direct internal conversion to the ground state via the S 1 /S 0 conical intersection, as well as intersystem crossing from T 1 to S 0 , appears to be less favorable or proceeds on a relatively slower timescale, primarily due to higher energetic barriers and the relatively weak SOC between T 1 and S 0 (41.6 cm − 1 ). As a result, the molecule is likely to remain in the T 1 state for an extended period, enabling it to engage in subsequent photochemical reactions. 3.2 [2 + 2] Cycloaddition from Tp 4 Se T to Se 5 -selenetane Minima and crossing points. As detailed above, the S 2 ( 1 ππ * ) excited state can undergo an efficient relaxation to the lowest triplet state, where the photoinduced [2 + 2] cycloaddition reaction is initiated. The triplet-state minima of Se 5 -selenetane, optimized at the QM(CASSCF)/MM level, are designated as T 1CC and T 1CSe in Fig. 6 , with the subscripts CC and CSe denoting the covalent bonding between C 4 –C 6’ and C 5’ –Se 8 , respectively. In both T 1CC and T 1CSe , only a single covalent bond is formed: C 4 –C 6’ (1.608 Å) in T 1CC and C 5’ –Se 8 (2.299 Å) in T 1CSe , while the other bond remains cleaved but exhibits a weak interaction. All attempts to optimize a T 1CCCSe structure, wherein both C 4 –C 6’ and C 5’ –Se 8 bonds are simultaneously formed in the triplet state, were unsuccessful. Similarly, optimization efforts targeting the corresponding ground-state structures—namely S 0CC , S 0CSe , and S 0CCCSe —were undertaken. Among these, only the S 0CCCSe structure was successfully located, characterized by the formation of both covalent bonds with bond lengths of 1.605 Å (C 4 –C 6’ ) and 1.980 Å (C 5’ –Se 8 ). The relative potential energies of T 1CC , T 1CSe , and S 0CCCSe are calculated to be 41.8, 48.7, and 6.1 kcal/mol, respectively. Table 5 QM(MS-CASPT2)/MM calculated relative energies (in kcal/mol) and spin-orbit couplings of minima and crossing points of Se5-selenetane. The potential energy of the S0 minimum of Tp4SeT is taken as the reference zero point. Structures Energies Structures Energies SOCs S 0CCCSe 6.1 S 0 /T 1CC 41.9/46.1 114.3 T 1CC 41.8 S 0 /T 1CSe 43.5/47.7 6.2 T 1CSe 48.7 Furthermore, two S 0 /T 1 crossing points, denoted as S 0 /T 1CC and S 0 /T 1CSe , were identified at the QM(CASSCF)/MM level. These crossings are anticipated to serve as key intermediates along the minimum-energy reaction pathways of the photoinduced [2 + 2] cycloaddition. As shown in Table 5 , the calculated relative energies and associated SOC constants are 41.9/46.1 kcal/mol [114.3 cm − 1 ] for S 0 /T 1CC and 43.5/47.7 kcal/mol [6.2 cm − 1 ] for S 0 /T 1CSe . Among these, the S 0 /T 1CC crossing point is predicted to be the most favorable for promoting the [2 + 2] cycloaddition, owing to its relatively moderate energetic barrier and substantially larger SOC value. [2 + 2] cycloaddition reaction. Due to the absence of a stable T 1CCCSe intermediate, the concerted pathway in the T 1 state is impossible and can be excluded from further consideration. In this respect, only two stepwise and nonadiabatic pathways via T 1CC and T 1CSe were considered for the photoinduced formation of Se 5 -selenetane from the Tp 4 Se T complex in the T 1 state (Fig. 7 ). In the first phase, the C 4 –C 6’ distance gradually decreases from 5.4 Å in the Tp 4 Se T dimer to 1.6 Å in the T 1CC intermediate. This process needs overcome an energy barrier of approximately 8 kcal/mol at the QM(MS-CASPT2//CASSCF)/MM level (Fig. 7 a). At this stage, the system approaches the S 0 /T 1CC crossing region and can rapidly relax to the ground state owing to the relatively large SOC value of 114.3 cm − 1 . Upon reaching the S 0 state, the system readily proceeds to form the [2 + 2] cycloaddition product of Se 5 -selenetane through a barrierless reaction pathway (Fig. 7 b). As shown in Fig. 7 c, the Tp 4 Se T system follows an alternative pathway via the T 1CSe intermediate. The formation of T 1CSe begins on a relatively flat potential energy surface and proceeds without an energy barrier. This is followed by intersystem crossing to the ground state near the S 0 /T 1CSe crossing point, where the SOC is calculated to be 6.2 cm − 1 . Once in the S 0 state, the formation of the C 4 –C 6’ bond becomes both barrierless and highly exothermic, rendering the entire reaction pathway not only energetically favorable but also structurally accessible (Fig. 7 d). Taken together, the stepwise and nonadiabatic pathways via T 1CC and T 1CSe are feasible routes for the [2 + 2] cycloaddition to form the final product of Se 5 -selenetane. 3.3 Rearrangement Reaction from Se 5 -selenetane to Se 5 -(6 − 4) Subsequently, the two-dimensional potential energy surface (2D-PES) associated with the transformation from the Se₅-selenetane intermediate to the final Se 5 -(6 − 4) product was explored at the QM(MS-CASPT2//CASSCF)/MM level. The ground-state (6 − 4) rearrangement is characterized as a concerted process, as evidenced by the 2D-PES analysis (indicated by the red arrow in Fig. 8 ). Importantly, the cleavage of the C 4 –Se 8 bond and the formation of the Se 8 –H 9 bond occur in a synchronous manner, ultimately leading to the generation of the Se₅-(6 − 4) product. This reaction proceeds with a relatively low activation barrier of approximately 21.5 kcal/mol in the ground state, a value readily surmountable given the excess energy derived from the S 2 Franck–Condon region. These results underscore the role of Se 5 -selenetane as a crucial intermediate in facilitating the formation of the (6 − 4) rearrangement product, Se 5 -(6 − 4). 3.4 Comparison with Tp Se 4 T. Next, a detailed comparative analysis of the photochemical reaction mechanisms of the 4-thiothymine–thymine (Tp 4 S T) and Tp 4 Se T dimers is presented. The maximum absorption band of Tp 4 Se T exhibits a pronounced red shift compared to that of Tp 4 S T in our calculations, with the vertical excitation energy of 93.2 kcal/mol for Tp 4 S T and 85.3 kcal/mol for Tp 4 Se T. The observed red shift permits Tp 4 Se T to harvest longer-wavelength photons with greater tissue penetration depth, thus improving its suitability for photodynamic therapeutic applications. Moreover, the energies of the multi-state intersection region involving S 2 , S 1 , T 2 , and T 1 are lower in the Tp 4 Se T system compared to Tp 4 S T. In Tp 4 Se T, the S 2 /S 1 /T 2 /T 1 intersection points lie at approximately 83.6, 80.9, 81.6, and 79.3 kcal/mol, respectively—slightly below its vertical excitation energy of 85.3 kcal/mol. In contrast, the corresponding intersection energies in Tp 4 S T are substantially higher—104.2, 100.9, 101.4, and 98.7 kcal/mol—relative to its vertical excitation energy of 93.2 kcal/mol. 48 This favorable energetic alignment in Tp 4 Se T significantly enhances the likelihood of efficient population of either the S 1 or T 1 state. Furthermore, selenium substitution markedly amplifies SOC in Tp 4 Se T due to its higher atomic mass. The S 1 -T 1 SOC reaches 425.0 cm − 1 , significantly exceeding the 94.1 cm − 1 observed in Tp 4 S T, thereby promoting more efficient ISC to the triplet manifold. Furthermore, selenium substitution appears to reduce the energy of the S 1 /S 0 conical intersection (70.4/66.2 kcal/mol) in the Tp 4 Se T system, suggesting a non-negligible likelihood of nonradiative relaxation to the ground state via this pathway. In contrast, this deactivation channel is energetically unfavorable and largely inaccessible in Tp 4 S T (S 1 /S 0 : 99.3/103.7 kcal/mol). In the [2 + 2] cycloaddition reaction, the selenium-substituted Tp 4 Se T system is unable to form the Se 5 -selenetane intermediate via concerted C–C and C–Se bond formation on the T 1 potential energy surface, as the resulting product cannot be stabilized in the triplet state. Nonetheless, Se 5 -selenetane can still be accessed via a stepwise and nonadiabatic relaxation pathway from the T 1 to the S 0 state through a C–C bond-forming S 0 /T 1 crossing, which is facilitated by a notably large SOC value of 114.3 cm − 1 . In contrast, the alternative C–Se bond-forming crossing exhibits a much smaller SOC value of only 6.2 cm − 1 , making this pathway kinetically less favorable. By comparison, the sulfur-substituted analogue of Tp 4 S T is capable of generating the S 5 -thietane product in the T 1 state; however, the concerted [2 + 2] reaction in this case is rendered inaccessible due to a prohibitively high activation barrier. 48 Although both C–C and C–S type S 0 /T 1 crossing points are energetically accessible in the sulfur-substituted system, the associated SOC values are below 5.0 cm − 1 , indicating that intersystem crossing through these channels is generally inefficient. In the context of the (6 − 4) rearrangement reaction, both Se 5 -selenetane and S 5 -thietane undergo concerted rearrangements on the ground-state potential energy surface to afford their respective (6 − 4) photoproducts. This transformation proceeds via a synchronous cleavage of the C 4 –S 8 (or C 4 –Se 8 ) bond and formation of the S 8 –H 9 (or Se 8 –H 9 ) bond. 4. Conclusion In this study, we employed the high-level QM(MS-CASPT2//CASSCF)/MM method to systematically investigate the nonadiabatic decay pathways leading to the T 1 state, followed by the subsequent [2 + 2] cycloaddition and (6 − 4) rearrangement reactions of the Tp 4 Se T dimer in aqueous solution (Fig. 9 ). Our calculations revealed five distinct nonadiabatic decay pathways from the initially excited S 2 state to the T 1 state. Paths I and II proceed via internal conversion through the S 2 /S 1 conical intersection, followed by ISC from S 1 to T 1 or T 2 , facilitated by significant SOCs and small energy gaps. Population in T 2 can further decay to T 1 through the T 2 /T 1 conical intersection. Paths IV and V involve either direct or indirect ISC through a four-state intersection (S 2 /S 1 /T 2 /T 1 ). These pathways are ultrafast and efficient, effectively driving the Tp 4 Se T system to the active T 1 state. In contrast, intersystem crossing from the T 1 state to the ground state via the S 0 /T 1 crossing, as well as internal conversion from S 1 to S 0 , are either inefficient or occur on a significantly longer timescale due to the relatively small SOC and high energy barriers. In addition, we also identified several energetically accessible [2 + 2] cycloaddition routes leading to the formation of Se 5 -selenetane, specifically the stepwise, nonadiabatic reactions on the T 1 surface via the T 1CC or T 1CSe intermediates (Fig. 7 ). Additionally, the (6 − 4) rearrangement from Se 5 -selenetane to Se 5 -(6 − 4) on the S 0 surface was explored. 2D-PES analysis (Fig. 8 ) reveals a concerted mechanism with a moderate barrier (~ 21.5 kcal/mol), which is surmountable with the excess energy from the S 2 Franck–Condon region. This supports the role of Se 5 -selenetane as a key intermediate in the formation of the (6 − 4) photoproduct. In summary, our results provide valuable mechanistic insights into the excited-state dynamics, [2 + 2] cycloaddition, and (6 − 4) rearrangement reactions of selenobase–nucleobase pairs in DNA. Declarations Competing interests The authors declare no competing interests. Author contributions Rui Zhao and Jia-Ling Dai: Methodology, Investigation, Data curation. Bo-Wen Yin and Shuai Li: Writing – original draft, Formal analysis. Bin-Bin Xie: Writing – review & editing, Supervision, Formal analysis. Acknowledgements This work was supported by the Key Science and Technology Project of Jinhua City (2023-1-093), the Zhejiang Provincial Natural Science Foundation of China (No. LZ23B030001), the National Key Research and Development Program of China (No. 2019YFA0709400). Data availability All relevant data are available from the authors upon request. 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Supplementary Files SIv2.docx Supporting Information for 'QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in an aqueous solution' SIv2b.docx Supporting Information for 'QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in an aqueous solution' Scheme1.png Scheme 1. The thymidine-4-selenothymidine (Tp 4Se T) dimer in aqueous solution studied in this work. Upon irradiation, Tp 4Se T first forms the four-membered Se 5 -selenetane intermediate, which then undergoes rearrangement to produce the open-ring Se 5 -(6-4) product. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6547757","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":454236992,"identity":"72cbcea2-8623-4d7a-9da5-6df9d6190ab9","order_by":0,"name":"Binbin Xie","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA6UlEQVRIiWNgGAWjYBACA2YGNoYEBgYeBvYGBgOw0AGitfAcIFYLA1ALGEgkQIUIaTFn5z324OGOwzL8ks8fFBe2Mcjx3Uhg/FyAR4tlM1+6QeKZNB7J2QkJxjPbGIwlbyQwS8/A57DDPGYSiW02PAa3Ew4Y87YxJG64kcDGzENYiwSP/c2DDSAt9cRqAdoiwcwA0pJgQEiLZTOPuUFiWxqPxJk0BmOecxKGM888bJbGp8Wc/4zZw59th+35248/M+Yps5HnO5588DM+LciADRhJEkCasYFIDQwMzA+IVjoKRsEoGAUjCgAAHYhCXZdPN5MAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0002-5347-2795","institution":"Zhejiang Normal University","correspondingAuthor":true,"prefix":"","firstName":"Binbin","middleName":"","lastName":"Xie","suffix":""},{"id":454236993,"identity":"d99f55f3-cbac-44bc-83d1-3cb88c881379","order_by":1,"name":"Zhao Rui","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Zhao","middleName":"","lastName":"Rui","suffix":""},{"id":454236994,"identity":"a134b605-bace-445d-9da2-85472faa4a8e","order_by":2,"name":"Jia-Ling Dai","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Jia-Ling","middleName":"","lastName":"Dai","suffix":""},{"id":454236995,"identity":"e165cb8e-acb0-4af5-9245-629cc860d938","order_by":3,"name":"Bo-Wen Yin","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Bo-Wen","middleName":"","lastName":"Yin","suffix":""},{"id":454236996,"identity":"673e794f-23f4-4552-af54-3560639aa083","order_by":4,"name":"Li Shuai","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Li","middleName":"","lastName":"Shuai","suffix":""}],"badges":[],"createdAt":"2025-04-28 12:16:58","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6547757/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6547757/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":82769873,"identity":"2b74dfdd-88cd-49f2-b68e-a1e202602e85","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":223953,"visible":true,"origin":"","legend":"\u003cp\u003eThe QM/MM system studied in this work: Tp\u003csup\u003e4Se\u003c/sup\u003eT was treated quantum mechanically, while the water molecules were modeled using molecular mechanics.\u003c/p\u003e","description":"","filename":"Picture2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/daa811b51507f26700c4794b.jpg"},{"id":82769876,"identity":"ed8c28ea-c097-44ec-9c16-7f6824e9dc44","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":162571,"visible":true,"origin":"","legend":"\u003cp\u003eQM(CASSCF)/MM optimized minima of Tp\u003csup\u003e4Se\u003c/sup\u003eT in the lowest six electronically singlet and triplet states (i.e., S\u003csub\u003e0\u003c/sub\u003e, S\u003csub\u003e1\u003c/sub\u003e, S\u003csub\u003e2\u003c/sub\u003e, T\u003csub\u003e1\u003c/sub\u003e, T\u003csub\u003e2 \u003c/sub\u003eand T\u003csub\u003e3\u003c/sub\u003e). Also shown are the selected bond lengths. See the ESI for Cartesian coordinates.\u003c/p\u003e","description":"","filename":"Picture3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/fe0d6518729f575fd2ecf58a.jpg"},{"id":82769874,"identity":"22bb0f78-de0e-4de7-aadf-7cf811d56fb2","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":71344,"visible":true,"origin":"","legend":"\u003cp\u003eQM(CASSCF)/MM-computed bond length differences (in Å) of the minima (a), conical intersections, and crossing points (b) of the 4- selenothymine chromophore relative to the counterparts of the S\u003csub\u003e0\u003c/sub\u003e minimum.\u003c/p\u003e","description":"","filename":"Picture4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/f7e9f22719ca237986153648.jpg"},{"id":82770625,"identity":"28c73123-50c0-4368-a4e1-c41d0039cba2","added_by":"auto","created_at":"2025-05-15 06:09:08","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":134737,"visible":true,"origin":"","legend":"\u003cp\u003eQM(CASSCF)/MM optimized conical intersections, and crossing points of Tp\u003csup\u003e4Se\u003c/sup\u003eT in the lowest five electronically singlet and triplet states (i.e., S\u003csub\u003e0\u003c/sub\u003e, S\u003csub\u003e1\u003c/sub\u003e, S\u003csub\u003e2\u003c/sub\u003e, T\u003csub\u003e1\u003c/sub\u003e and T\u003csub\u003e2\u003c/sub\u003e). Also shown are the selected bond lengths. See the ESI for Cartesian coordinates.\u003c/p\u003e","description":"","filename":"Picture5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/c10dc358e8bfd59df07f1213.jpg"},{"id":82769890,"identity":"141e842a-87de-4549-8672-d7e5ac54188d","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":155201,"visible":true,"origin":"","legend":"\u003cp\u003eQM(MS-CASPT2//CASSCF)/MM computed LIIC paths of Tp\u003csup\u003e4Se\u003c/sup\u003eT connecting (left) the S\u003csub\u003e2\u003c/sub\u003e Franck–Condon point to the conical intersection S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e; (right) the S\u003csub\u003e2\u003c/sub\u003e Franck–Condon point to the crossing point S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e. Values in parentheses are energies, and values in square brackets are SOCs.\u003c/p\u003e","description":"","filename":"Picture6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/17e923e2e838eafbae8e7a6f.jpg"},{"id":82771104,"identity":"b1be0515-ca88-48a3-bf43-f32382e5141d","added_by":"auto","created_at":"2025-05-15 06:17:08","extension":"jpg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":135857,"visible":true,"origin":"","legend":"\u003cp\u003eQM(CASSCF)/MM optimized minima and crossing points of Se\u003csup\u003e5\u003c/sup\u003e-selenetane in the S\u003csub\u003e0\u003c/sub\u003e and T\u003csub\u003e1\u003c/sub\u003e states, where the subscript CC refers to the covalently bonded C\u003csub\u003e4\u003c/sub\u003e–C\u003csub\u003e6’\u003c/sub\u003e and CSe represents the C\u003csub\u003e5’\u003c/sub\u003e–Se\u003csub\u003e8\u003c/sub\u003e bond. Also shown are selected bond lengths. See the ESI for Cartesian coordinates.\u003c/p\u003e","description":"","filename":"Picture7.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/6f809ba3a092e3174cb1b4fe.jpg"},{"id":82769880,"identity":"130f4ebc-beb4-4878-9707-9d21a27a83c0","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"jpg","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":158934,"visible":true,"origin":"","legend":"\u003cp\u003eQM(MS-CASPT2//CASSCF)/MM optimized minimum-energy reaction paths from Tp\u003csup\u003e4Se\u003c/sup\u003eT to Se\u003csup\u003e5\u003c/sup\u003e-selenetane in the T\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e0\u003c/sub\u003e states via the T\u003csub\u003e1CC\u003c/sub\u003e (a-b) and T\u003csub\u003e1CSe\u003c/sub\u003e (c-d) intermediates. The subscript CC refers to the covalently bonded C\u003csub\u003e4\u003c/sub\u003e–C\u003csub\u003e6’\u003c/sub\u003e and CSe represents the C\u003csub\u003e5’\u003c/sub\u003e–Se\u003csub\u003e8\u003c/sub\u003e bond.\u003c/p\u003e","description":"","filename":"Picture8.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/76774364730fb6717838ae9b.jpg"},{"id":82769887,"identity":"2f8013ac-18f9-4f4c-90f3-4cf8494a8da6","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"jpg","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":138766,"visible":true,"origin":"","legend":"\u003cp\u003eQM(MS-CASPT2//CASSCF)/MM optimized two-dimensional potential energy surface from Se\u003csup\u003e5\u003c/sup\u003e-selenetane to Se\u003csup\u003e5\u003c/sup\u003e-(6-4) in the S\u003csub\u003e0\u003c/sub\u003e state with geometric constraints on the C\u003csub\u003e4\u003c/sub\u003e–Se\u003csub\u003e8\u003c/sub\u003e and Se\u003csub\u003e8\u003c/sub\u003e–H\u003csub\u003e9\u003c/sub\u003e bonds (energies in kcal/mol).\u003c/p\u003e","description":"","filename":"Picture9.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/f93d33b6eb6c6b7e0d477e38.jpg"},{"id":82769881,"identity":"e03e9bce-01cd-4d1f-a581-21287e75d2d7","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"jpg","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":98751,"visible":true,"origin":"","legend":"\u003cp\u003eSuggested photophysical processes and photochemical reaction mechanisms based on the present computational results\u003cstrong\u003e.\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Picture10.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/bb7be2e0fa8ebfd80c38e66d.jpg"},{"id":84555692,"identity":"400e1a57-c018-4209-ae9b-687a725002c8","added_by":"auto","created_at":"2025-06-13 11:28:45","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2529515,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/5a076347-4e39-47f1-aad3-abee7a73d5c7.pdf"},{"id":82769879,"identity":"92249249-7e41-4701-a6cd-cd19e6a7710b","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":906215,"visible":true,"origin":"","legend":"Supporting Information for 'QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in an aqueous solution'","description":"","filename":"SIv2.docx","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/1a781babcc8dd186ecf68a6e.docx"},{"id":82769883,"identity":"5eca494f-964e-43a1-bb98-0956e4aba487","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"docx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":905891,"visible":true,"origin":"","legend":"Supporting Information for 'QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in an aqueous solution'","description":"","filename":"SIv2b.docx","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/51a02734f2d7cafdaa67c7b8.docx"},{"id":82769878,"identity":"d765d714-4003-4c7e-a008-bda76651bd53","added_by":"auto","created_at":"2025-05-15 06:01:08","extension":"png","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":65756,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 1. \u003c/strong\u003eThe thymidine-4-selenothymidine (Tp\u003csup\u003e4Se\u003c/sup\u003eT) dimer in aqueous solution studied in this work. Upon irradiation, Tp\u003csup\u003e4Se\u003c/sup\u003eT first forms the four-membered Se\u003csup\u003e5\u003c/sup\u003e-selenetane intermediate, which then undergoes rearrangement to produce the open-ring Se\u003csup\u003e5\u003c/sup\u003e-(6-4) product.\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e","description":"","filename":"Scheme1.png","url":"https://assets-eu.researchsquare.com/files/rs-6547757/v1/db65595a0719e35803a89b0c.png"}],"financialInterests":"There is \u003cb\u003eNO\u003c/b\u003e Competing Interest.","formattedTitle":"QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in aqueous solution","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003e \u003c/p\u003e \u003cp\u003eAmong various phototherapeutic approaches, photodynamic therapy has emerged as a clinically approved and effective modality that utilizes light to activate photosensitizers (PSs) \u0026mdash; photoactive compounds capable of producing reactive oxygen species (ROS).\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e,\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e Upon photoexcitation to their triplet states, PSs mediate ROS production through two primary pathways: Type I mechanisms involve electron/hydrogen transfer with biomolecules to form superoxide anions (O\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e) or hydroxyl radicals (\u0026middot;OH), while Type II mechanisms utilize triplet-triplet energy transfer to convert ground-state oxygen (\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003eO\u003csub\u003e2\u003c/sub\u003e) into cytotoxic singlet oxygen (\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003eO\u003csub\u003e2\u003c/sub\u003e). These ROS species play pivotal roles in selective apoptosis induction for therapeutic applications spanning antimicrobial treatment, dermatological disorders, and oncological interventions.\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e,\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e Current clinically approved PSs predominantly comprise porphyrin derivatives and transition metal complexes, where metal incorporation enhances triplet quantum yields. However, challenges persist regarding oxygen dependency (particularly problematic in hypoxic tumor microenvironments) and non-specific phototoxicity toward healthy tissues.\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e,\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e Addressing these limitations necessitates rational design of novel PSs with optimized ROS generation efficiency, hypoxia tolerance, and biological selectivity \u0026mdash; goals requiring fundamental understanding of photophysical processes (e.g., excited-state lifetimes, intersystem crossing efficiency) and photochemical reactions within biological contexts.\u003c/p\u003e \u003cp\u003eTo circumvent metal-induced cytotoxicity, recent efforts have focused on developing metal-free PS systems.\u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e,\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e Sulfur-substituted PSs have emerged as promising candidates due to their tunable photophysics.\u003csup\u003e\u003cspan additionalcitationids=\"CR10\" citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e Strategic substitution of carbonyl oxygen with sulfur atoms in molecular frameworks enables enhanced intersystem crossing (ISC) and near-unity triplet quantum yields. In particular, thionucleobases have garnered significant attention as photosensitizers owing to their structural resemblance to natural nucleobases, along with several advantageous properties: (1) excellent biocompatibility, (2) red-shifted absorption profiles that facilitate deeper tissue penetration, (3) large photon absorption cross-sections, and (4) strong spin-orbit coupling combined with small singlet\u0026ndash;triplet energy gaps, enabling highly efficient triplet-state population.\u003csup\u003e\u003cspan additionalcitationids=\"CR13 CR14 CR15 CR16 CR17 CR18 CR19 CR20 CR21 CR22 CR23 CR24 CR25 CR26\" citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eExpanding this strategy to heavier chalcogens, selenonucleobases have recently gained experimental and theoretical interest.\u003csup\u003e\u003cspan additionalcitationids=\"CR29 CR30 CR31 CR32 CR33 CR34 CR35 CR36 CR37 CR38 CR39 CR40 CR41 CR42 CR43 CR44\" citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e\u003c/sup\u003e Crespo-Hern\u0026aacute;ndez and colleagues utilized time-resolved spectroscopy to probe the photochemical properties of 6-thioguanine and its selenium analogue, 6-selenoguanine.\u003csup\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e\u003c/sup\u003e Their comparative analysis revealed that replacing sulfur with selenium leads to a further red-shift in absorption and promotes ISC, though it also results in a relatively short triplet-state lifetime. From a theoretical perspective, Pirillo et al. conducted DFT and TD-DFT computations to evaluate the photophysical characteristics, including absorption profiles, excited-state energies, and spin\u0026ndash;orbit coupling elements, of thymidine and deoxyguanosine substituted with selenium and tellurium.\u003csup\u003e\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e,\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e\u003c/sup\u003e Their findings indicate that these compounds exhibit efficient ISC and possess adequate triplet energies to activate molecular oxygen, suggesting their promise as UVA-targeted therapeutic agents. Additionally, Hazra and Manae investigated the influence of electronic conjugation and chalcogen atomic size on the spectral and reactive properties of modified thymine molecules.\u003csup\u003e\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e\u003c/sup\u003e In parallel, Mai and collaborators carried out multireference electronic structure calculations and nonadiabatic dynamics simulations at the MS-CASPT2 and ADC(2) levels, unveiling rapid population and depopulation processes of the reactive triplet state in 2-selenouracil.\u003csup\u003e\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e More recently, Fang\u0026rsquo;s group applied MS-CASPT2 and combined QM(MS-CASPT2)/MM methods to examine the excited-state relaxation mechanisms of 6-selenoguanine in different environments, including gas phase, aqueous solution, and DNA matrices.\u003csup\u003e\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e,\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e Besides, Peng et al. systematically explored the photophysical behavior of 2-selenouracil, 4-selenouracil, and 2,4-diselenouracil through MS-CASPT2 calculations, highlighting the position-dependent influence of selenium substitution on excited-state dynamics.\u003csup\u003e\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e\u003c/sup\u003e Furthermore, Valverde et al. used QM/MM dynamics to study 6-selenoguanine in water and found an ISC time of 452 fs, longer than the experimental 130 fs, which reflects multiple overlapping processes rather than ISC alone.\u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eWhile these studies have offered valuable mechanistic insights into selenonucleobases, current researches have primarily focused on their electronic structures and nonadiabatic dynamics related to photophysical behavior. In contrast, their photochemical reactions remain insufficiently characterized. To fill this gap, we employed high-level complete active space self-consistent field (CASSCF) and its multistate second-order perturbation extension (MS-CASPT2) with in the quantum mechanics/molecular mechanics (QM/MM) framework to investigate the excited-state relaxation pathways, [2\u0026thinsp;+\u0026thinsp;2] photocycloaddition, and subsequent (6\u0026thinsp;\u0026minus;\u0026thinsp;4) photoproduct formation of the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT dimer in aqueous solution (Scheme \u003cspan refid=\"Sch1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). In addition, we further compared these systems with thymine:4-thiothymine (Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT) to assess selenium substitution effects on both photophysical and photochemical behaviors of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT.\u003csup\u003e\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e"},{"header":"2. Computational details","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 System setup\u003c/h2\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTo account for solvation effects, a spherical water box (radius\u0026thinsp;=\u0026thinsp;25 \u0026Aring;) containing 1977 water molecules was generated using Packmol.\u003csup\u003e\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e\u003c/sup\u003e The Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT dimer was placed at the center of this box. A quick energy minimization (2000 steps) was performed to eliminate unphysical molecular contacts, followed by a 50 ps MD simulation at 300 K using a N\u0026oacute;se\u0026ndash;Hoover thermostat. During the simulation, water molecules beyond 20 \u0026Aring; from the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT center of mass were frozen. The Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT dimer was modeled using the all-atom CHARMM general force field,\u003csup\u003e\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e,\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e\u003c/sup\u003e while the solvent molecules were described using the TIP3P model.\u003csup\u003e\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e\u003c/sup\u003e The final MD snapshot was used to prepare the QM/MM system by removing fixed waters, resulting in a 20 \u0026Aring; solvation shell with 3144 atoms (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). All simulations were conducted using CHARMM (version c31b1).\u003csup\u003e\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 QM/MM calculations\u003c/h2\u003e \u003cp\u003eFollowing equilibration MD simulations, all subsequent electronic structure calculations, including geometry optimizations and single-point energy refinements, were performed using the high-level complete active space self-consistent field (CASSCF) and its multistate second-order perturbation extension (MS-CASPT2) with in the QM/MM framework.\u003csup\u003e\u003cspan additionalcitationids=\"CR55\" citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e\u003c/sup\u003e In this hybrid scheme, the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT dimer (30 atoms) was treated quantum mechanically at MS-CASPT2//CASSCF level, while the MM subsystem employed the TIP3P water model.\u003csup\u003e\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e\u003c/sup\u003e The simulation system consisted of 2694 mobile atoms (within 15 \u0026Aring; of the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT dimer) and 450 fixed peripheral atoms. The electronic structure calculations employed an active space comprising sixteen electrons distributed across twelve orbitals for both CASSCF and MS-CASPT2 treatments (\u003cb\u003eFigure \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e\u003c/b\u003e). Key computational parameters included: (1) Cholesky decomposition with unbiased auxiliary basis sets for efficient two-electron integral evaluation;\u003csup\u003e\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e\u003c/sup\u003e (2) zero ionization potential-electron affinity (IPEA) shift;\u003csup\u003e\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e\u003c/sup\u003e and (3) an imaginary shift of 0.2 atomic units to mitigate intruder-state artifacts.\u003csup\u003e\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e\u003c/sup\u003e Spin-orbit coupling (SOC) constants were computed using the atomic mean-field approximation (AMFI) approach:\u003csup\u003e\u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e\u003c/sup\u003e\u003cdiv id=\"Equa\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equa\" name=\"EquationSource\"\u003e\n$$\\:⟨{\\psi\\:}_{I}\\left|{H}_{eff}^{SOC}\\right|{\\psi\\:}_{J}⟩\\:=\\:\\sqrt{\\:[⟨{\\psi\\:}_{I}|{H}_{x}^{SOC}|{\\psi\\:}_{J}⟩\u0026sup2;\\:+\\:⟨{\\psi\\:}_{I}|{H}_{y}^{SOC}|{\\psi\\:}_{J}⟩\u0026sup2;\\:+\\:⟨{\\psi\\:}_{I}|{H}_{z}^{SOC}|{\\psi\\:}_{J}⟩\u0026sup2;]\\:/\\:3}\\:$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eAll quantum mechanical calculations, including both QM(CASSCF)/MM and QM(MS-CASPT2)/MM approaches, were carried out using the aug-cc-pVDZ basis set for the tellurium atom and the cc-pVDZ basis set for all other atoms.\u003csup\u003e\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e,\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e\u003c/sup\u003e These calculations were implemented through the OpenMolcas program (version v21.02)\u003csup\u003e\u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e\u003c/sup\u003e in conjunction with the TINKER molecular modeling package (version 6.3.2).\u003csup\u003e\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results and Discussion","content":"\u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e3.1 Excited-State Decay of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT\u003c/h2\u003e \u003cp\u003e \u003cb\u003eVertical excitation properties.\u003c/b\u003e We initially performed geometry optimization for the ground-state minimum of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT at the QM(CASSCF)/MM level, which is denoted as S\u003csub\u003e0\u003c/sub\u003e in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e. On this optimized structure, vertical excitation energies and electronic configurations of the low-lying excited states, namely, S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003en\u003csub\u003eH\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*), S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eπ\u003csub\u003eH\u0026minus;1\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*), T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eπ\u003csub\u003eH\u0026minus;1\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*), T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003en\u003csub\u003eH\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*), and T\u003csub\u003e3\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eπ\u003csub\u003eH\u0026minus;4\u003c/sub\u003eπ\u003csub\u003eL\u003c/sub\u003e*), were examined based on QM(MS-CASPT2)/MM calculations (see Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eFrontier molecular orbital analysis indicates that these excitations are predominantly localized on the 4-selenothymine moiety. Specifically, the HOMO corresponds to a nonbonding n orbital centered on the Se atom (Se\u003csub\u003e8\u003c/sub\u003e), the HOMO-1 is attributed to the π orbital of the C\u003csub\u003e4\u003c/sub\u003e\u0026thinsp;=\u0026thinsp;Se\u003csub\u003e8\u003c/sub\u003e bond, and the LUMO\u0026thinsp;+\u0026thinsp;1 is a delocalized π* orbital spanning the 4-selenothymine ring (\u003cb\u003eFigure \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e\u003c/b\u003e). The S\u003csub\u003e1\u003c/sub\u003e state arises primarily from a HOMO \u0026rarr; LUMO\u0026thinsp;+\u0026thinsp;1 transition, with a dominant configuration weight of 0.56. As expected, this state is optically forbidden with a negligible oscillator strength. The QM(MS-CASPT2)/MM method predicted vertical excitation energy of 58.5 kcal/mol for the S\u003csub\u003e1\u003c/sub\u003e state, which is 5.0 kcal/mol lower than the value reported by Pirillo et al.\u003csup\u003e\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/sup\u003e In contrast, the S\u003csub\u003e2\u003c/sub\u003e state is optically allowed, involving a major HOMO-1 \u0026rarr; LUMO\u0026thinsp;+\u0026thinsp;1 transition (weight: 0.67), and exhibits a significantly higher oscillator strength. The computed vertical excitation energy for the S₂ state (85.3 kcal/mol) at the Franck\u0026ndash;Condon geometry in a water environment is consistent with the value reported by Pirillo et al., who obtained 83.0 kcal/mol for 4-selenothymidine in water using the IEFPCM model.\u003csup\u003e\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/sup\u003e Additionally, Shi et al. experimentally determined the maximum absorption band of 4-selenothymidine to be 77.7 kcal/mol using UV spectroscopy, which is slightly lower than the values predicted by theoretical calculations.\u003csup\u003e\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e This discrepancy may be attributed to the fact that, as noted by G\u0026ouml;tze et al., vertical excitation energies are typically higher than experimental absorption maxima, since the latter reflect transitions from vibrationally relaxed states.\u003csup\u003e\u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eAdditionally, the vertical excitation energies for the T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eπ\u003csub\u003eH\u0026minus;1\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*) and T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003en\u003csub\u003eH\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*) states were calculated to be 55.4 and 56.6 kcal/mol, respectively, which are in good agreement with the values reported by Pirillo et al. at the M06/6\u0026ndash;31\u0026thinsp;+\u0026thinsp;G* level.\u003csup\u003e46\u003c/sup\u003e The electronic configurations of T\u003csub\u003e1\u003c/sub\u003e and T\u003csub\u003e2\u003c/sub\u003e correspond to ππ* and nπ* characters, respectively. The T\u003csub\u003e3\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eπ\u003csub\u003eH\u0026minus;4\u003c/sub\u003eπ\u003csub\u003eL\u003c/sub\u003e*) state exhibits a significantly higher vertical excitation energy\u0026mdash;approximately 5.0 kcal/mol above that of the S\u003csub\u003e2\u003c/sub\u003e state. This might indicate that the T\u003csub\u003e3\u003c/sub\u003e state plays a relatively minor role during the excited-state relaxation process.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eQM(MS-CASPT2//CASSCF)/MM computed vertical excitation energies (kcal/mol) of the lowest several excited singlet and triplet states of Tp4SeT.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStates\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eMS-CASPT2\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eM06/6\u0026ndash;31\u0026thinsp;+\u0026thinsp;G*\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eExp.\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003en\u003csub\u003eH\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e58.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e63.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eπ\u003csub\u003eH\u0026minus;1\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e85.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e83.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e77.7\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eπ\u003csub\u003eH\u0026minus;1\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e55.4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e50.7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003en\u003csub\u003eH\u003c/sub\u003eπ\u003csub\u003eL+1\u003c/sub\u003e*)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e56.6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e58.6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT\u003csub\u003e3\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eπ\u003csub\u003eH\u0026minus;4\u003c/sub\u003eπ\u003csub\u003eL\u003c/sub\u003e*)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e90.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026mdash;\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"4\" nameend=\"c4\" namest=\"c1\"\u003e \u003cp\u003e\u003csup\u003ea\u003c/sup\u003e Theor. Chem. Acc. 2016, 135, 8; \u003csup\u003eb\u003c/sup\u003e Nucleos. Nucleot. Nucl. 2021, 40, 96\u0026ndash;116.\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eAdiabatic excitation properties.\u003c/b\u003e \u003c/p\u003e \u003cp\u003eAs illustrated in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e, we optimized the minimum-energy geometries of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT in the S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e), S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e), T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e), T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e), and T\u003csub\u003e3\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) states at the QM(CASSCF)/MM level without imposing any geometric constraints. The electronic transitions and associated structural rearrangements are mainly localized on the 4-selenothymine chromophore, except for the T\u003csub\u003e3\u003c/sub\u003e minimum, where the excitation is primarily centered on the thymine unit. Figure\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e presents the bond length differences between these five excited-state minima and the ground-state minimum.\u003c/p\u003e \u003cp\u003eThe most significant structural changes are observed in the C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e, C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e5\u003c/sub\u003e, and C\u003csub\u003e5\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u003c/sub\u003e bond lengths, while variations in other bonds remain relatively minor compared to the S\u003csub\u003e0\u003c/sub\u003e structure. For example, in the S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) minimum, the C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e bond is markedly elongated to 1.989 \u0026Aring;, approximately 0.2 \u0026Aring; longer than in the S\u003csub\u003e0\u003c/sub\u003e minimum. Additionally, the C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e5\u003c/sub\u003e bond shortens by over 0.12 \u0026Aring;, while the C\u003csub\u003e5\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u003c/sub\u003e bond lengthens by more than 0.11 \u0026Aring;. The structural variations in the S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e), T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e), and T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) minima exhibit similar trends relative to S\u003csub\u003e0\u003c/sub\u003e, with bond length variations generally confined within the range of \u0026minus;\u0026thinsp;0.12 to 0.12 \u0026Aring;.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe adiabatic excitation energies corresponding to the T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e), T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e), T\u003csub\u003e3\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e), S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e), and S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) minima were computed to be 46.1, 49.6, 76.4, 48.4, and 66.2 kcal/mol, respectively, at the QM(MS-CASPT2)/MM level (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Notably, at the S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) minimum geometry, the single-point energies of the S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e), T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e), and T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) states are nearly degenerate, with values of 48.4, 48.1, and 47.6 kcal/mol, respectively. This near-degeneracy implies that the S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) minimum could serve as a key intersection point in the excited-state relaxation pathways. In contrast, the relatively high adiabatic excitation energy associated with the T\u003csub\u003e3\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) minimum suggests that this state is less likely to contribute significantly to the excited-state decay dynamics.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eQM(MS-CASPT2//CASSCF)/MM calculated relative energies (in kcal/mol) of minima of Tp4SeT. The potential energy of the S0 minimum is taken as the reference zero point.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStructures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eEnergies\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStructures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eEnergies\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e0\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eT\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e46.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e48.4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eT\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e49.6\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e66.2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eT\u003csub\u003e3\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e76.4\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eSeveral surface-crossing structures involving the S\u003csub\u003e0\u003c/sub\u003e, S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e), S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e), T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e), and T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) states of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT were identified through QM(CASSCF)/MM calculations. These include three minimum-energy conical intersections between singlet (S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e) and triplet (T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e) states, along with four singlet\u0026ndash;triplet crossing points (S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e, S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e, S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e, and S\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e). The optimized geometries and corresponding bond length variations for these conical intersections and crossing points are depicted in Figs.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e and \u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e.\u003c/p\u003e \u003cp\u003eNearly all intersection structures exhibit substantial geometric distortions compared to the S\u003csub\u003e0\u003c/sub\u003e minimum. In particular, pronounced bond length changes, ranging from \u0026minus;\u0026thinsp;0.16 to 0.35 \u0026Aring;, are observed for key bonds such as C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e, N\u003csub\u003e3\u003c/sub\u003e\u0026ndash;C\u003csub\u003e4\u003c/sub\u003e, C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e5\u003c/sub\u003e, and C\u003csub\u003e5\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u003c/sub\u003e. Interestingly, only the S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e, intersection structures retain an essentially planar configuration. By contrast, the T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e conical intersections show slight out-of-plane displacement of the Se\u003csub\u003e8\u003c/sub\u003e atom relative to the pyrimidine ring. Moreover, prominent pyramidalization of carbon or nitrogen atoms is observed in the S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e, S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e, and S\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e crossings, indicating strong out-of-plane distortions associated with the surface-crossing geometries.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eEnergetically, the S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e conical intersections are located at 83.6/80.9 and 70.4/66.2 kcal/mol, respectively. Given that the vertical excitation energy of S\u003csub\u003e2\u003c/sub\u003e is 85.3 kcal/mol, both intersections are readily accessible, thus promoting efficient internal conversion pathways back to the ground state. Notably, at the S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e conical intersection, the S\u003csub\u003e1\u003c/sub\u003e, S\u003csub\u003e2\u003c/sub\u003e, T\u003csub\u003e1\u003c/sub\u003e and T\u003csub\u003e2\u003c/sub\u003e states are nearly degenerate, forming a unique four-state crossing. This feature may facilitate both internal conversion and intersystem crossing. The T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e conical intersection lies at a relatively low energy (51.8/50.2 kcal/mol), suggesting that rapid internal conversion from T\u003csub\u003e2\u003c/sub\u003e to T\u003csub\u003e1\u003c/sub\u003e to is likely to occur through this intersection.\u003c/p\u003e \u003cp\u003eRegarding the crossing points, the single-point energies of S\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e are calculated to be 89.7/88.6 kcal/mol, approximately 4.0 kcal/mol higher than the vertical excitation energy at the S\u003csub\u003e2\u003c/sub\u003e Franck\u0026ndash;Condon point. This indicates that access to the S\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e crossing is energetically unfavorable, and thus it likely plays a minor role in the excited-state deactivation processes. In contrast, the S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e crossing structures can be regarded as three-state intersections (S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e), with single-point energies of 57.5/53.9/53.2 and 54.8/54.2/54.2 kcal/mol, respectively. As expected, the S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e crossing structure exhibits a relatively higher energy of 67.1/68.3 kcal/mol. This observation aligns with the experimentally reported high triplet yield in the overall excited-state relaxation dynamics.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eQM(MS-CASPT2//CASSCF)/MM calculated relative energies (in kcal/mol) of conical intersections and crossing points of Tp4SeT. The potential energy of the S0 minimum is taken as the reference zero point.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStructures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eEnergies\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStructures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eEnergies\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e70.4/66.2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e54.8/54.2\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e83.6/80.9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e57.5/53.9\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e51.8/50.2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e89.7/88.6\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e67.1/68.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"4\" nameend=\"c4\" namest=\"c1\"\u003e \u003cp\u003e\u003csup\u003ea\u003c/sup\u003e S₂/S₁ forms a four-state crossing (S\u003csub\u003e1\u003c/sub\u003e, S\u003csub\u003e2\u003c/sub\u003e, T\u003csub\u003e1\u003c/sub\u003e and T\u003csub\u003e2\u003c/sub\u003e) with close energies: 83.6, 80.9, 81.6, and 79.3 kcal/mol; \u003csup\u003eb\u003c/sup\u003e S₁/T₁ and S₁/T₂ are three-state crossings (S\u003csub\u003e1\u003c/sub\u003e, T\u003csub\u003e1\u003c/sub\u003e and T\u003csub\u003e2\u003c/sub\u003e) with energies of 54.8/54.2/54.2 and 57.5/53.9/53.2 kcal/mol.\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eExcited-state decay pathways.\u003c/b\u003e \u003c/p\u003e \u003cp\u003eTo gain further insight into the efficient triplet-state population in Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT, we performed linearly interpolated internal coordinate (LIIC) path calculations at the QM(MS-CASPT2)/MM level of theory (Figs.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e and S2). Starting from the S\u003csub\u003e2\u003c/sub\u003e Franck\u0026ndash;Condon region, five potential nonadiabatic decay pathways were identified along the LIIC profiles: Path I: S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e)-FC \u0026rarr; S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e \u0026rarr; S\u003csub\u003e1\u003c/sub\u003e \u0026rarr; S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e \u0026rarr; T\u003csub\u003e1\u003c/sub\u003e;Path II: S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e)-FC \u0026rarr; S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e \u0026rarr; S\u003csub\u003e1\u003c/sub\u003e \u0026rarr; S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e \u0026rarr; T\u003csub\u003e2\u003c/sub\u003e \u0026rarr; T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e \u0026rarr; T\u003csub\u003e1\u003c/sub\u003e; Path III: S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e)-FC \u0026rarr; S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e \u0026rarr; S\u003csub\u003e1\u003c/sub\u003e \u0026rarr; S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e \u0026rarr; S\u003csub\u003e0\u003c/sub\u003e. Path IV: S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e)-FC \u0026rarr; S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e \u0026rarr; T\u003csub\u003e1\u003c/sub\u003e; Path V: S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e)-FC \u0026rarr; S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e \u0026rarr; T\u003csub\u003e2\u003c/sub\u003e \u0026rarr; T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e \u0026rarr; T\u003csub\u003e1\u003c/sub\u003e.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eQM(MS-CASPT2)/MM calculated spin-orbit couplings (in cm\u0026minus;\u0026thinsp;1) at several important intersection structures of Tp4SeT.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"7\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStructures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eStates\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSOCs\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eStructures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eStates\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eSOCs\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eSOCs\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e-T\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e306.6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eS\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eS\u003csub\u003e0\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e41.6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e\u0026mdash;\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e160.6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e425.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e891.5\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e162.9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e17.8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e17.3\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eS\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e394.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eS\u003csub\u003e2\u003c/sub\u003e-T\u003csub\u003e2\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e346.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e303.7\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colspan=\"7\" nameend=\"c7\" namest=\"c1\"\u003e \u003cp\u003e\u003csup\u003ea\u003c/sup\u003e Theor. Chem. Acc. 2016, 135, 8.\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eWe begin by discussing the excited-state decay pathway that involves only internal conversion, namely Path III. As shown in the left panel of Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e, this pathway requires overcoming an energy barrier of approximately 14.0 kcal/mol from the S\u003csub\u003e2\u003c/sub\u003e minimum to the S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e conical intersection. The relative energy at this conical intersection (83.6/80.9 kcal/mol) remains below that of the S\u003csub\u003e2\u003c/sub\u003e Franck\u0026ndash;Condon region (85.3 kcal/mol), indicating that the initially populated S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) state can undergo internal conversion to the S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) state near this region due to the small energy gap. Subsequently, the S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) population can decay to the ground state via the S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e conical intersection, although this process requires overcoming an energy barrier of approximately 20 kcal/mol. The right panel of Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e illustrates a similar initial process, where the population transitions from the initially excited S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) state to the S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) state via the S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e conical intersection. Unlike Path I, the decay pathways of Paths II and III are more likely to proceed via intersystem crossing to the T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) or T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) states through the S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e or S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e crossings. This is facilitated by their near-degeneracy and the significant SOCs of 425.0 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e (S\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e) and 17.8 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e (S\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e2\u003c/sub\u003e). Subsequently, the T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) state can further relax to the lower-lying T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) state via the T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e conical intersection.\u003c/p\u003e \u003cp\u003eAs illustrated in \u003cb\u003eFigure S2\u003c/b\u003e, transitions from S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) to the reactive T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) state via the four-state intersection of S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e are also accessible (Paths IV and V). These processes benefit from both efficient internal conversion and intersystem crossing. At this multistate intersection, the molecule may directly transition to T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) (Path IV), aided by a S\u003csub\u003e2\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e SOC of 160.6 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, or proceed indirectly via intermediate T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) state with a S\u003csub\u003e2\u003c/sub\u003e-T\u003csub\u003e2\u003c/sub\u003e SOC of 306.6 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e (Path V). From this intermediate, further relaxation to T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) is highly probable due to T\u003csub\u003e2\u003c/sub\u003e \u0026rarr; T\u003csub\u003e1\u003c/sub\u003e internal conversion.\u003c/p\u003e \u003cp\u003eThe proposed excited-state decay pathways above are consistent with El-Sayed's rule,\u003csup\u003e\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e\u003c/sup\u003e which states that ISC is significantly enhanced when it involves a change in orbital character. For instance, the SOC constants between S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) and T\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) at the S\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e crossing, and between S\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003enπ\u003csup\u003e*\u003c/sup\u003e) and T\u003csub\u003e1\u003c/sub\u003e(\u003csup\u003e3\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) at the S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e crossing were estimated to be 346.0 and 425.0 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, respectively. Notably, at the S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e multi-state intersection, the electronic configurations of both singlet (S\u003csub\u003e2\u003c/sub\u003e, S\u003csub\u003e1\u003c/sub\u003e) and triplet (T\u003csub\u003e2\u003c/sub\u003e, T\u003csub\u003e1\u003c/sub\u003e) states exhibit strong mixing of nπ\u003csup\u003e*\u003c/sup\u003e and ππ\u003csup\u003e*\u003c/sup\u003e characters. This leads to exceptionally large SOCs not only for S\u003csub\u003e2\u003c/sub\u003e-T\u003csub\u003e2\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e, but also for S\u003csub\u003e2\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e2\u003c/sub\u003e transitions.\u003c/p\u003e \u003cp\u003eTaking into account both the relative energetics and the computed SOC constants, we concluded that the ultrafast and energetically accessible nonadiabatic decay pathways efficiently promote triplet-state population in Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT. In contrast, direct internal conversion to the ground state via the S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e conical intersection, as well as intersystem crossing from T\u003csub\u003e1\u003c/sub\u003e to S\u003csub\u003e0\u003c/sub\u003e, appears to be less favorable or proceeds on a relatively slower timescale, primarily due to higher energetic barriers and the relatively weak SOC between T\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e0\u003c/sub\u003e (41.6 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e). As a result, the molecule is likely to remain in the T\u003csub\u003e1\u003c/sub\u003e state for an extended period, enabling it to engage in subsequent photochemical reactions.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e3.2 [2\u0026thinsp;+\u0026thinsp;2] Cycloaddition from Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT to Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane\u003c/h2\u003e \u003cp\u003e \u003cb\u003eMinima and crossing points.\u003c/b\u003e \u003c/p\u003e \u003cp\u003eAs detailed above, the S\u003csub\u003e2\u003c/sub\u003e(\u003csup\u003e1\u003c/sup\u003eππ\u003csup\u003e*\u003c/sup\u003e) excited state can undergo an efficient relaxation to the lowest triplet state, where the photoinduced [2\u0026thinsp;+\u0026thinsp;2] cycloaddition reaction is initiated. The triplet-state minima of Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane, optimized at the QM(CASSCF)/MM level, are designated as T\u003csub\u003e1CC\u003c/sub\u003e and T\u003csub\u003e1CSe\u003c/sub\u003e in Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e, with the subscripts CC and CSe denoting the covalent bonding between C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u0026rsquo;\u003c/sub\u003e and C\u003csub\u003e5\u0026rsquo;\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e, respectively. In both T\u003csub\u003e1CC\u003c/sub\u003e and T\u003csub\u003e1CSe\u003c/sub\u003e, only a single covalent bond is formed: C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u0026rsquo;\u003c/sub\u003e (1.608 \u0026Aring;) in T\u003csub\u003e1CC\u003c/sub\u003e and C\u003csub\u003e5\u0026rsquo;\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e (2.299 \u0026Aring;) in T\u003csub\u003e1CSe\u003c/sub\u003e, while the other bond remains cleaved but exhibits a weak interaction. All attempts to optimize a T\u003csub\u003e1CCCSe\u003c/sub\u003e structure, wherein both C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u0026rsquo;\u003c/sub\u003e and C\u003csub\u003e5\u0026rsquo;\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e bonds are simultaneously formed in the triplet state, were unsuccessful. Similarly, optimization efforts targeting the corresponding ground-state structures\u0026mdash;namely S\u003csub\u003e0CC\u003c/sub\u003e, S\u003csub\u003e0CSe\u003c/sub\u003e, and S\u003csub\u003e0CCCSe\u003c/sub\u003e\u0026mdash;were undertaken. Among these, only the S\u003csub\u003e0CCCSe\u003c/sub\u003e structure was successfully located, characterized by the formation of both covalent bonds with bond lengths of 1.605 \u0026Aring; (C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u0026rsquo;\u003c/sub\u003e) and 1.980 \u0026Aring; (C\u003csub\u003e5\u0026rsquo;\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e). The relative potential energies of T\u003csub\u003e1CC\u003c/sub\u003e, T\u003csub\u003e1CSe\u003c/sub\u003e, and S\u003csub\u003e0CCCSe\u003c/sub\u003e are calculated to be 41.8, 48.7, and 6.1 kcal/mol, respectively.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eQM(MS-CASPT2)/MM calculated relative energies (in kcal/mol) and spin-orbit couplings of minima and crossing points of Se5-selenetane. The potential energy of the S0 minimum of Tp4SeT is taken as the reference zero point.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStructures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eEnergies\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eStructures\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eEnergies\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eSOCs\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eS\u003csub\u003e0CCCSe\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e6.1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eS\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CC\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e41.9/46.1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e114.3\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT\u003csub\u003e1CC\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e41.8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eS\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CSe\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e43.5/47.7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e6.2\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eT\u003csub\u003e1CSe\u003c/sub\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e48.7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eFurthermore, two S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e crossing points, denoted as S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CC\u003c/sub\u003e and S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CSe\u003c/sub\u003e, were identified at the QM(CASSCF)/MM level. These crossings are anticipated to serve as key intermediates along the minimum-energy reaction pathways of the photoinduced [2\u0026thinsp;+\u0026thinsp;2] cycloaddition. As shown in Table\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e5\u003c/span\u003e, the calculated relative energies and associated SOC constants are 41.9/46.1 kcal/mol [114.3 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e] for S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CC\u003c/sub\u003e and 43.5/47.7 kcal/mol [6.2 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e] for S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CSe\u003c/sub\u003e. Among these, the S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CC\u003c/sub\u003e crossing point is predicted to be the most favorable for promoting the [2\u0026thinsp;+\u0026thinsp;2] cycloaddition, owing to its relatively moderate energetic barrier and substantially larger SOC value.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003e[2\u0026thinsp;+\u0026thinsp;2] cycloaddition reaction.\u003c/b\u003e \u003c/p\u003e \u003cp\u003eDue to the absence of a stable T\u003csub\u003e1CCCSe\u003c/sub\u003e intermediate, the concerted pathway in the T\u003csub\u003e1\u003c/sub\u003e state is impossible and can be excluded from further consideration. In this respect, only two stepwise and nonadiabatic pathways via T\u003csub\u003e1CC\u003c/sub\u003e and T\u003csub\u003e1CSe\u003c/sub\u003e were considered for the photoinduced formation of Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane from the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT complex in the T\u003csub\u003e1\u003c/sub\u003e state (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn the first phase, the C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u0026rsquo;\u003c/sub\u003e distance gradually decreases from 5.4 \u0026Aring; in the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT dimer to 1.6 \u0026Aring; in the T\u003csub\u003e1CC\u003c/sub\u003e intermediate. This process needs overcome an energy barrier of approximately 8 kcal/mol at the QM(MS-CASPT2//CASSCF)/MM level (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ea). At this stage, the system approaches the S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CC\u003c/sub\u003e crossing region and can rapidly relax to the ground state owing to the relatively large SOC value of 114.3 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e. Upon reaching the S\u003csub\u003e0\u003c/sub\u003e state, the system readily proceeds to form the [2\u0026thinsp;+\u0026thinsp;2] cycloaddition product of Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane through a barrierless reaction pathway (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003eb).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eAs shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ec, the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT system follows an alternative pathway via the T\u003csub\u003e1CSe\u003c/sub\u003e intermediate. The formation of T\u003csub\u003e1CSe\u003c/sub\u003e begins on a relatively flat potential energy surface and proceeds without an energy barrier. This is followed by intersystem crossing to the ground state near the S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1CSe\u003c/sub\u003e crossing point, where the SOC is calculated to be 6.2 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e. Once in the S\u003csub\u003e0\u003c/sub\u003e state, the formation of the C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;C\u003csub\u003e6\u0026rsquo;\u003c/sub\u003e bond becomes both barrierless and highly exothermic, rendering the entire reaction pathway not only energetically favorable but also structurally accessible (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003ed). Taken together, the stepwise and nonadiabatic pathways via T\u003csub\u003e1CC\u003c/sub\u003e and T\u003csub\u003e1CSe\u003c/sub\u003e are feasible routes for the [2\u0026thinsp;+\u0026thinsp;2] cycloaddition to form the final product of Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e3.3 Rearrangement Reaction from Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane to Se\u003csup\u003e5\u003c/sup\u003e-(6\u0026thinsp;\u0026minus;\u0026thinsp;4)\u003c/h2\u003e \u003cp\u003eSubsequently, the two-dimensional potential energy surface (2D-PES) associated with the transformation from the Se₅-selenetane intermediate to the final Se\u003csup\u003e5\u003c/sup\u003e-(6\u0026thinsp;\u0026minus;\u0026thinsp;4) product was explored at the QM(MS-CASPT2//CASSCF)/MM level. The ground-state (6\u0026thinsp;\u0026minus;\u0026thinsp;4) rearrangement is characterized as a concerted process, as evidenced by the 2D-PES analysis (indicated by the red arrow in Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e). Importantly, the cleavage of the C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e bond and the formation of the Se\u003csub\u003e8\u003c/sub\u003e\u0026ndash;H\u003csub\u003e9\u003c/sub\u003e bond occur in a synchronous manner, ultimately leading to the generation of the Se₅-(6\u0026thinsp;\u0026minus;\u0026thinsp;4) product. This reaction proceeds with a relatively low activation barrier of approximately 21.5 kcal/mol in the ground state, a value readily surmountable given the excess energy derived from the S\u003csub\u003e2\u003c/sub\u003e Franck\u0026ndash;Condon region. These results underscore the role of Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane as a crucial intermediate in facilitating the formation of the (6\u0026thinsp;\u0026minus;\u0026thinsp;4) rearrangement product, Se\u003csup\u003e5\u003c/sup\u003e-(6\u0026thinsp;\u0026minus;\u0026thinsp;4).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e3.4 Comparison with Tp\u003csup\u003eSe\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003eT.\u003c/h2\u003e \u003cp\u003eNext, a detailed comparative analysis of the photochemical reaction mechanisms of the 4-thiothymine\u0026ndash;thymine (Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT) and Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT dimers is presented.\u003c/p\u003e \u003cp\u003eThe maximum absorption band of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT exhibits a pronounced red shift compared to that of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT in our calculations, with the vertical excitation energy of 93.2 kcal/mol for Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT and 85.3 kcal/mol for Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT. The observed red shift permits Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT to harvest longer-wavelength photons with greater tissue penetration depth, thus improving its suitability for photodynamic therapeutic applications. Moreover, the energies of the multi-state intersection region involving S\u003csub\u003e2\u003c/sub\u003e, S\u003csub\u003e1\u003c/sub\u003e, T\u003csub\u003e2\u003c/sub\u003e, and T\u003csub\u003e1\u003c/sub\u003e are lower in the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT system compared to Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT. In Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT, the S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e intersection points lie at approximately 83.6, 80.9, 81.6, and 79.3 kcal/mol, respectively\u0026mdash;slightly below its vertical excitation energy of 85.3 kcal/mol. In contrast, the corresponding intersection energies in Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT are substantially higher\u0026mdash;104.2, 100.9, 101.4, and 98.7 kcal/mol\u0026mdash;relative to its vertical excitation energy of 93.2 kcal/mol.\u003csup\u003e48\u003c/sup\u003e This favorable energetic alignment in Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT significantly enhances the likelihood of efficient population of either the S\u003csub\u003e1\u003c/sub\u003e or T\u003csub\u003e1\u003c/sub\u003e state. Furthermore, selenium substitution markedly amplifies SOC in Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT due to its higher atomic mass. The S\u003csub\u003e1\u003c/sub\u003e-T\u003csub\u003e1\u003c/sub\u003e SOC reaches 425.0 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, significantly exceeding the 94.1 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e observed in Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT, thereby promoting more efficient ISC to the triplet manifold. Furthermore, selenium substitution appears to reduce the energy of the S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e conical intersection (70.4/66.2 kcal/mol) in the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT system, suggesting a non-negligible likelihood of nonradiative relaxation to the ground state via this pathway. In contrast, this deactivation channel is energetically unfavorable and largely inaccessible in Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT (S\u003csub\u003e1\u003c/sub\u003e/S\u003csub\u003e0\u003c/sub\u003e: 99.3/103.7 kcal/mol).\u003c/p\u003e \u003cp\u003eIn the [2\u0026thinsp;+\u0026thinsp;2] cycloaddition reaction, the selenium-substituted Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT system is unable to form the Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane intermediate via concerted C\u0026ndash;C and C\u0026ndash;Se bond formation on the T\u003csub\u003e1\u003c/sub\u003e potential energy surface, as the resulting product cannot be stabilized in the triplet state. Nonetheless, Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane can still be accessed via a stepwise and nonadiabatic relaxation pathway from the T\u003csub\u003e1\u003c/sub\u003e to the S\u003csub\u003e0\u003c/sub\u003e state through a C\u0026ndash;C bond-forming S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e crossing, which is facilitated by a notably large SOC value of 114.3 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e. In contrast, the alternative C\u0026ndash;Se bond-forming crossing exhibits a much smaller SOC value of only 6.2 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, making this pathway kinetically less favorable. By comparison, the sulfur-substituted analogue of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eS\u003c/sup\u003eT is capable of generating the S\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-thietane product in the T\u003csub\u003e1\u003c/sub\u003e state; however, the concerted [2\u0026thinsp;+\u0026thinsp;2] reaction in this case is rendered inaccessible due to a prohibitively high activation barrier.\u003csup\u003e\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e\u003c/sup\u003e Although both C\u0026ndash;C and C\u0026ndash;S type S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e crossing points are energetically accessible in the sulfur-substituted system, the associated SOC values are below 5.0 cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, indicating that intersystem crossing through these channels is generally inefficient.\u003c/p\u003e \u003cp\u003eIn the context of the (6\u0026thinsp;\u0026minus;\u0026thinsp;4) rearrangement reaction, both Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane and S\u003csub\u003e5\u003c/sub\u003e-thietane undergo concerted rearrangements on the ground-state potential energy surface to afford their respective (6\u0026thinsp;\u0026minus;\u0026thinsp;4) photoproducts. This transformation proceeds via a synchronous cleavage of the C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;S\u003csub\u003e8\u003c/sub\u003e (or C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e) bond and formation of the S\u003csub\u003e8\u003c/sub\u003e\u0026ndash;H\u003csub\u003e9\u003c/sub\u003e (or Se\u003csub\u003e8\u003c/sub\u003e\u0026ndash;H\u003csub\u003e9\u003c/sub\u003e) bond.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"4. Conclusion","content":"\u003cp\u003eIn this study, we employed the high-level QM(MS-CASPT2//CASSCF)/MM method to systematically investigate the nonadiabatic decay pathways leading to the T\u003csub\u003e1\u003c/sub\u003e state, followed by the subsequent [2\u0026thinsp;+\u0026thinsp;2] cycloaddition and (6\u0026thinsp;\u0026minus;\u0026thinsp;4) rearrangement reactions of the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT dimer in aqueous solution (Fig.\u0026nbsp;\u003cspan refid=\"Fig9\" class=\"InternalRef\"\u003e9\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eOur calculations revealed five distinct nonadiabatic decay pathways from the initially excited S\u003csub\u003e2\u003c/sub\u003e state to the T\u003csub\u003e1\u003c/sub\u003e state. Paths I and II proceed via internal conversion through the S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e conical intersection, followed by ISC from S\u003csub\u003e1\u003c/sub\u003e to T\u003csub\u003e1\u003c/sub\u003e or T\u003csub\u003e2\u003c/sub\u003e, facilitated by significant SOCs and small energy gaps. Population in T\u003csub\u003e2\u003c/sub\u003e can further decay to T\u003csub\u003e1\u003c/sub\u003e through the T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e conical intersection. Paths IV and V involve either direct or indirect ISC through a four-state intersection (S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e). These pathways are ultrafast and efficient, effectively driving the Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT system to the active T\u003csub\u003e1\u003c/sub\u003e state. In contrast, intersystem crossing from the T\u003csub\u003e1\u003c/sub\u003e state to the ground state via the S\u003csub\u003e0\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e crossing, as well as internal conversion from S\u003csub\u003e1\u003c/sub\u003e to S\u003csub\u003e0\u003c/sub\u003e, are either inefficient or occur on a significantly longer timescale due to the relatively small SOC and high energy barriers.\u003c/p\u003e \u003cp\u003eIn addition, we also identified several energetically accessible [2\u0026thinsp;+\u0026thinsp;2] cycloaddition routes leading to the formation of Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane, specifically the stepwise, nonadiabatic reactions on the T\u003csub\u003e1\u003c/sub\u003e surface via the T\u003csub\u003e1CC\u003c/sub\u003e or T\u003csub\u003e1CSe\u003c/sub\u003e intermediates (Fig.\u0026nbsp;\u003cspan refid=\"Fig7\" class=\"InternalRef\"\u003e7\u003c/span\u003e). Additionally, the (6\u0026thinsp;\u0026minus;\u0026thinsp;4) rearrangement from Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane to Se\u003csup\u003e5\u003c/sup\u003e-(6\u0026thinsp;\u0026minus;\u0026thinsp;4) on the S\u003csub\u003e0\u003c/sub\u003e surface was explored. 2D-PES analysis (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e8\u003c/span\u003e) reveals a concerted mechanism with a moderate barrier (~\u0026thinsp;21.5 kcal/mol), which is surmountable with the excess energy from the S\u003csub\u003e2\u003c/sub\u003e Franck\u0026ndash;Condon region. This supports the role of Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane as a key intermediate in the formation of the (6\u0026thinsp;\u0026minus;\u0026thinsp;4) photoproduct.\u003c/p\u003e \u003cp\u003eIn summary, our results provide valuable mechanistic insights into the excited-state dynamics, [2\u0026thinsp;+\u0026thinsp;2] cycloaddition, and (6\u0026thinsp;\u0026minus;\u0026thinsp;4) rearrangement reactions of selenobase\u0026ndash;nucleobase pairs in DNA.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e \u003ch2\u003eCompeting interests\u003c/h2\u003e \u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e \u003c/p\u003e\u003ch2\u003eAuthor contributions\u003c/h2\u003e \u003cp\u003eRui Zhao and Jia-Ling Dai: Methodology, Investigation, Data curation. Bo-Wen Yin and Shuai Li: Writing \u0026ndash; original draft, Formal analysis. Bin-Bin Xie: Writing \u0026ndash; review \u0026amp; editing, Supervision, Formal analysis.\u003c/p\u003e\u003ch2\u003eAcknowledgements\u003c/h2\u003e \u003cp\u003eThis work was supported by the Key Science and Technology Project of Jinhua City (2023-1-093), the Zhejiang Provincial Natural Science Foundation of China (No. LZ23B030001), the National Key Research and Development Program of China (No. 2019YFA0709400).\u003c/p\u003e\u003ch2\u003eData availability\u003c/h2\u003e \u003cp\u003eAll relevant data are available from the authors upon request. Rui Zhao and Bin-Bin Xie will be responsible for replying to this request. A complete set of original computational files, comprising input files, output files, and molden files, will be provided in a ZIP archive.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003ePham, T. C., Nguyen, V.-N., Choi, Y., Lee, S. \u0026amp; Yoon, J. 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Res. 1, 8\u0026ndash;16 (1968).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"},{"header":"Scheme ","content":"\u003cp\u003eScheme 1 is available in the Supplementary Files section.\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-6547757/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6547757/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eSelenonucleobases have garnered increasing interest from both experimental and theoretical communities for their promising roles in photodynamic therapy and DNA crosslinking. Similar to the extensively investigated thymidine:4-thiothymidine system, the selenium-modified thymidine:4-selenothymidine (Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT) dimer may also exhibit significant photochemical activity within DNA duplexes. However, its detailed photochemical reaction mechanisms remain largely unexplored. Herein, we employed high-level MS-QM(CASPT2//CASSCF) method to explore excited-state decay, [2\u0026thinsp;+\u0026thinsp;2] cycloaddition and (6\u0026thinsp;\u0026minus;\u0026thinsp;4) reactions of Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT in aqueous solution. Our calculations revealed five possible nonadiabatic decay channels enabling population of the T\u003csub\u003e1\u003c/sub\u003e state from the initial S\u003csub\u003e2\u003c/sub\u003e state, mediated by two multi-state intersections of S\u003csub\u003e2\u003c/sub\u003e/S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e and S\u003csub\u003e1\u003c/sub\u003e/T\u003csub\u003e2\u003c/sub\u003e/T\u003csub\u003e1\u003c/sub\u003e. Following population of the T\u003csub\u003e1\u003c/sub\u003e state, the [2\u0026thinsp;+\u0026thinsp;2] cycloaddition proceeds via a stepwise, nonadiabatic mechanism. That is, the pathway starts from Tp\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003eSe\u003c/sup\u003eT in the T\u003csub\u003e1\u003c/sub\u003e state via the T\u003csub\u003e1CC\u003c/sub\u003e or T\u003csub\u003e1CSe\u003c/sub\u003e intermediates and ultimately ends up with Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane in the S\u003csub\u003e0\u003c/sub\u003e state. Subsequent transformation of Se\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e-selenetane into the Se\u003csup\u003e5\u003c/sup\u003e-(6\u0026thinsp;\u0026minus;\u0026thinsp;4) product occurs through a concerted reaction in the ground state, characterized by the simultaneous cleavage of the C\u003csub\u003e4\u003c/sub\u003e\u0026ndash;Se\u003csub\u003e8\u003c/sub\u003e bond and formation of the S\u003csub\u003e8\u003c/sub\u003e\u0026ndash;H\u003csub\u003e9\u003c/sub\u003e bond. This study provides detailed mechanistic insights into the photoreactivity of selenonucleobases in DNA duplexes at the molecular level.\u003c/p\u003e","manuscriptTitle":"QM/MM studies on photo-induced cycloaddition and (6-4) reactions of the thymidine:4-selenothymidine dimer in aqueous solution","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-05-15 06:01:03","doi":"10.21203/rs.3.rs-6547757/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"dfe4dd0e-8404-4752-b40f-5ead066e077f","owner":[],"postedDate":"May 15th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":48301614,"name":"Physical sciences/Chemistry/Theoretical chemistry/Computational chemistry"},{"id":48301615,"name":"Physical sciences/Chemistry/Physical chemistry/Excited states"},{"id":48301616,"name":"Physical sciences/Chemistry/Photochemistry/Photobiology"}],"tags":[],"updatedAt":"2025-06-13T11:20:37+00:00","versionOfRecord":[],"versionCreatedAt":"2025-05-15 06:01:03","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6547757","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6547757","identity":"rs-6547757","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}