Electronic Relaxation Pathways in Heavy-Atom-Free Photosensitizers Absorbing Near-Infrared Radiation and Exhibiting High Yields of Singlet Oxygen Generation

Abstract

Heavy-atom-free photosensitizers (HAF-PSs) based on thionation of carbonyl groups of readily accessible organic compounds are rapidly emerging as a versatile class of molecules. However, their photochemical properties and electronic relaxation mechanisms are currently unknown. Investigating the excited-state dynamics is essential to understand their benefits and limitations and to develop photosensitizers with improved photochemical properties. Herein, the photochemical and electronic-structure properties of two of the most promising HAF-PSs developed to date are revealed. It is shown that excitation of thio-4-(dimethylamino)naphthalamide and thionated Nile Red with near-infrared radiation leads to the efficient population of the triplet manifold through multiple relaxation pathways in hundreds of femtoseconds. The strong singlet–triplet couplings in this family of photosensitizers should enable a broad range of applications, including in photodynamic therapy, photocatalysis, photovoltaics, organic LEDs, and photon up-conversion.

Photodynamic therapy (PDT) is a clinically approved, noninvasive therapy for cancer treatment that relies on the administration of a photosensitizer (PS) and light to the affected area.^1-5 Photoactivation of the PS promotes intersystem crossing (ISC) to the triplet state, which can transfer energy to molecular oxygen, generating singlet oxygen and other reactive oxygen species (ROS) that are toxic to the targeted cancer cells and tissues. PDT is more attractive than conventional therapies such as radiotherapy and chemotherapy, because it is minimally invasive, has exceptional spatiotemporal selectivity and diminished side effects, and has an overall simplicity among other important considerations.^4,6-8 Notwithstanding the significant benefits over traditional therapies, PDT has yet to reach its full potential primarily because optimal PSs, applicable to a wide range of cancers and biological tissues, are difficult to develop. An ideal PS should be easy to synthesize and store,⁹ has a high molar absorption in the therapeutic window (ca. 600–800 nm),¹⁰ high triplet (Φ_T) and singlet oxygen (Φ_Δ) quantum yields, and low cytotoxicity in the dark, and be cost-effective.^5,9,11

One prevalent approach is to incorporate heavy atoms into organic molecules to enhance their ISC efficiency, yet these compounds are often difficult to synthesize, costly, and suffer from increased dark cytotoxicity.^12-14 Another approach that is swiftly gaining increased attention is to thionate carbonyl groups of existing organic molecules to red shift and increase their absorption coefficients and increase their Φ_T and Φ_Δ.^5,15-18 Many of these heavy-atom-free PSs (HAF-PSs), offer good biocompatibility, biodegradability, minimal dark cytotoxicity, and structural stability.^5,16,18-20 Two highly promising HAF-PSs are thio-4-(dimethylamino)naphthalamide (SDMNP) and thionated Nile Red (SNR).^18,21 These HAF-PSs exhibit high near-infrared (NIR) cross sections, high Φ_Δ, and good PDT efficacy against monolayer cancer (HeLa) cells and 3D multicellular tumor spheroids.¹⁸ However, their photochemical properties are currently unknown, particularly, their electronic relaxation mechanisms upon NIR excitation. Elucidating the relaxation mechanisms of these HAF-PSs is essential for diagnosing their benefits and limitations, and to refine this chemistry.

Figure 1 shows the absorption spectra of SDMNP and SNR in DMSO and MeCN. The spectrum of SDMNP extends to 725 and 750 nm in MeCN and DMSO, respectively, while it extends to 750 and 795 nm for SNR. Both molecules exhibit negligible fluorescence quantum yields (<0.001) and generate high yields of singlet oxygen (81% and 36% for SDMNP and SNR, respectively).^18,21

Figure 1.

Absorption spectra of (a) SDMNP and (b) SNR in dimethyl sulfoxide (DMSO) and acetonitrile (MeCN). Vertical lines correspond to the calculated electronic transitions and corresponding oscillator strengths at the TD-PBE0/IEFPCM/6-31+G(d,p)//B3LYP/IEFPCM/6-31+G(d,p) in each solvent. Inset: molecular structure of SDMNP and SNR.

Vertical excitation energies (VEEs) were calculated to characterize the accessible electronic transitions for ISC to reactive triplet states upon NIR excitation (see Figure 1 and Tables S1 and S2 for SDMNP and SNR, respectively). The S₁ state has nπ* character and negligible oscillator strength in both molecules, which rationalizes their negligible fluorescence yields instead of photoinduced electron transfer,^18,21 while the S₂ state has ππ* character and is directly populated upon NIR excitation. There are four and three triplet states lower in energy than the S₂(ππ*) state in SDMNP and SNR, respectively. Hence, the population reaching the S₂(ππ*) state could relax from the Franck–Condon (FC) region through at least five electronic states in SDMNP or through four in SNR. Furthermore, within the expected accuracy of the calculations (±0.3 eV), the S₂(ππ*) state is isoenergetic with the S₁(nπ*) and the T₄(nπ*) states in SDMNP, while the S₁(nπ*) state is isoenergetic with the T₃(ππ*) state in SNR, suggesting that these states are strongly coupled in the FC region, possibly giving rise to efficient ISC to the triplet manifold.

The spin–orbit coupling constants (SOCs) between the two lowest-energy excited singlet states and the four/three triplet states of SDMNP/SNR were also calculated at three key regions of their potential energy surfaces (PESs): the FC, the S₁ minimum, and the S₂ minimum (Tables S3 and S4). On the basis of the SOCs and El Sayed’s rules,^22,23 S₂ → T₃ and S₁ → T₁ are predicted to occur more efficiently in SDMNP, with S₂ → T₄ playing a smaller role, while S₂ → T₂, S₁ → T₁, and S₁ → T₃ are predicted to occur more efficiently in SNR. The efficiency of these pathways in both molecules is independent of the regions of the PESs or solvent used. The multitude of ISC pathways predicted to play a role is remarkable, confirming the increased density of states upon thionation,²⁴ which can give rise to efficient and near-unity Φ_T.

Femtosecond transient absorption spectroscopy (fs-TAS) was used to investigate if ISC occurs efficiently and independent of solvent. Figure 2 presents the TAS for SDMNP following 610 nm excitation. Two positive- and two negative-amplitude bands are observed within the pump–probe cross-correlation (Figure 2a,d). The negative-amplitude bands are assigned to S₀ depopulation, in agreement with the absorption spectra (Figure 1a). The positive-amplitude bands are assigned to excited-state absorption (ESA) by transient species. One of the bands exhibits a maximum around 340 nm, while the other has a maximum around 460 nm. Both bands reach their maximum intensity around 240 and 350 fs in DMSO and MeCN, respectively. A blue shift of the 460 nm band is observed (Figure 2b,e), while a new absorption band appears simultaneously with a maximum around 680 nm. The positive-amplitude absorption bands decay monotonically during ca. 11 ps to 3 ns in DMSO, or ca. 30 ps to 3 ns in MeCN, while S₀ repopulation is simultaneously observed. Representative decay traces, lifetimes, and corresponding evolution-associated difference spectra (EADS) are reported in Figure 3a,b, Table S5, and Figure S1, respectively.

Figure 2.

fs-TAS for SDMNP (λ_exc = 610 nm) in (a)–(c) DMSO and (d)–(f) MeCN solutions. Solvent-stimulated Raman emission bands are observed within the pump–probe cross-correlation around 510 and 520 nm for DMSO and MeCN, respectively. The breaks are covering the scattering of the pump reaching the detectors.

Figure 3.

Representative decay traces for SDMNP in (a) DMSO and (b) MeCN and for SNR in (c) DMSO and (d) MeCN. The solid lines are the fittings obtained a kinetic analysis of the transient absorption data (see Methods in the Supporting Information for details).

To assist in the assignment of SDMNP’s transient species, the S₂(ππ*), S₁(nπ*), and T₁(ππ*) states were optimized and their ESAs were calculated (Figure S2). Attempts to optimize the T₃(nπ*) state were unsuccessful. The ESA demonstrates that the absorption spectra of both singlet states and the T₁(ππ*) state overlap significantly, while the ESA from the singlet excited states are largely blue-shifted relative to that of the T₁(ππ*) state. Hence, we assign the 460 nm absorption band to the vibrationally excited T₁(ππ*) and/or T₃(nπ*) states, with some overlap with the S₂(ππ*) and S₁(nπ*) states. The 339 nm band has significant overlap from both S₁(nπ*) and S₂(ππ*) states. A comparison of decay traces at 339 and 460 nm support this assignment (Figure 3a,b). The band between 400 and 525 nm red shifts and grows within the pump–probe cross-correlation (Figures 2a,d and 3). The red shift is assigned to the population of the S₂(ππ*) state and ultrafast internal conversion (IC) to the S₁(nπ*) state, competing with ISC to the triplet manifold from both singlet states. Indeed, considering El Sayed’s rules, the large SOCs, and the dynamics observed in the TAS, it becomes evident that population of the triplet manifold occurs through two competitive pathways: (1) a direct S₂(ππ*) → T₃(nπ*) pathway, with subsequent IC to the T₁(ππ*) state, and (2) an indirect S₂(ππ*) → S₁(nπ*) → T₁(ππ*) ISC pathway. Remarkably, both pathways are occurring within 580 and 210 fs (τ₁) in DMSO and MeCN, respectively. The ultrafast lifetime suggests that the triplet state of SDMNP is populated in high yield, as observed for a wide-range of thiobase derivatives^15,16,24-33 and other thionated compounds.^34-36

The blue shift observed in Figure 2b,e is assigned to a combination of vibrational cooling (VC) and solvent relaxation (SR) dynamics in the triplet manifold, occurring within 5 ps in DMSO and 50 ps in MeCN. The large ΔE(S₁–T₁) and ΔE(T₃–T₁) energy gaps (Table S1) and the observation that τ₂ changes significantly with solvent (Table S5) support this assignment. The optimized structures suggest that blue shifting is not due to major conformational changes in the excited states (Figure S3).

The population reaching the triplet state of SDMNP decays within 3 ns in DMSO and 60% slower in MeCN (Figure 2c,f). ISC back to S₀ occurs rapidly, suggesting that there is significant SOC between the T₁ and S₀ states. Triplet decay is not due to triplet self-quenching in the concentration from 75 to 779 μM (Figure S4), suggesting that it is due to an intrinsic process. Fast, hundreds of picoseconds ISC back to S₀ has been observed previously for a selenium-substituted DNA base.³⁷ Scheme 1a depicts the proposed relaxation mechanism.

Scheme 1. Proposed Deactivation Mechanism for SDMNP (a) and SNR (b) in DMSO and MeCN^a.

^aThickness of the arrow represents qualitatively the probability of a given pathway (the thicker the arrow, the higher the probability).

Figure 4 presents the TAS for SNR following 674 nm excitation. Positive- and negative-amplitude bands are observed within the pump–probe cross-correlation (Figure 4a,b). The negative-amplitude bands are assigned to S₀ depopulation, in agreement with the absorption spectra (Figure 1b). Three positive-amplitude absorption bands are observed with maxima around 405, 450, and 550 nm. A minor absorption band is observed around 330 nm in both solvents, but it strongly overlaps with the S₀-depopulation signal. The 550 nm band starts to decay during the initial hundreds of femtoseconds in DMSO (Figure 3b), while the other bands stay relatively unchanged. In MeCN, all bands start to decay during the initial hundreds of femtoseconds (Figure 4f). As the 550 nm band continues to decay and shifts to 570 nm (Figure 4c), the other two bands start to decay as well (Figure 4f,g). Additionally, the 448 nm band blue shifts (Figure 4c, inset). Figure 4d,g shows that the residual transient signal decays monotonically, but not fully within 3 ns. Representative decay traces, lifetimes, and corresponding EADS are reported in Figure 3c,d, Table S5, and Figure S5, respectively.

Figure 4.

fs-TAS for SNR (λ_exc = 674 nm) in (a)–(d) DMSO and (e)–(g) MeCN solutions. The solvent-stimulated Raman emission band is observed at 555 nm. The breaks are covering the scattering of the pump reaching the detectors.

To assist in the assignment of the SNR’s transient species, the S₂(ππ*), S₁(nπ*), and T₁(ππ*) states were optimized and their ESA spectra calculated. Attempts to optimize the T₂(nπ*) and T₃(ππ) states were unsuccessful. On the basis of the energy gaps (Table S2), SOCs (Table S5), and calculated ESA spectra (Figure S6), the absorption bands at 330 and 448 nm in Figure 4a,e are assigned to a combination of the S₁(nπ*) and S₂(ππ*) states, while the bands at 405 and 550–570 nm are assigned to a triplet state (i.e., T₂ and/or T₃), the vibrationally excited T₁(ππ*) state, or a combination of them. This is evident in Figure 3c,d, where the kinetic traces at 448 nm rapidly decay at an early time, following a decay behavior similar to that of the species absorbing at 405 nm. Hence, the complex dynamics observed during the first few picoseconds in Figure 4 correspond to a competition between ultrafast S₂(ππ*) → S₁(nπ*) IC and ISC from both singlet states to the triplet manifold (i.e., T₃, T₂, and/or T₁). This combination of processes occurs within 500 fs (τ₁), independent of solvent. As for SDMNP, VC and SR in the triplet manifold occur within τ₂, which is supported by the change of τ₂ with solvent (Table S5). The dynamics in Figure 4d,g correspond to the partial decay of the T₁(ππ*) state together with S₀ repopulation. Scheme 1b depicts the proposed relaxation mechanism.

In this study, the deactivation mechanisms of two promising HAF-PSs¹⁸ were elucidated and the primary doorway states leading to ultrafast ISC dynamics were identified. Thionation of carbonyl groups is key to enable the photochemical properties exhibited by these HAF-PSs. VEEs and SOCs for dimethylaminonaphtalamide and Nile Red are reported in Tables S6-S9, which provide full support and strengthen the notion that thionation is a convenient and general strategy to red shift the absorption spectrum, increase the density of states, and the singlet–triplet SOCs of organic compounds, enabling ultrafast ISC, and high Φ_T and Φ_Δ.^5,16,24,32 Besides PDT, HAF-PSs are anticipated to find wide-ranging applications in photocatalysis, photovoltaics, organic LEDs, and photon up-conversion.