Measuring the Efficiency of Raman Photoexcitation of Singlet Oxygen in Distilled Water

1. Introduction

Singlet oxygen (¹O₂) corresponds to the lowest excited electronic quantum level of molecular oxygen. In this state, the molecule is highly electrophilic, which is the basis for its broad applications in photochemistry, photobiology, and photomedicine [1-5]. Quantum mechanics forbids direct one-photon excitation toward the ¹O₂ state, resulting in a low transition probability. Photosensitization is the general method for efficient ¹O₂ photoproduction [6-7]. A large photosensitizer molecule absorbs visible (VI) light, accumulating the absorbed energy on a triple metastable level. From there, the energy is transmitted to surrounding oxygen molecules, exciting the ¹O₂ state. We have recently demonstrated a photosensitizer-free ¹O₂ photoexcitation based on the Raman effect [8-10]. A pumping photon produces a Raman transition from ground toward the ¹O₂ level, emitting a Stokes photon of lower energy. The difference between the pumping and Stokes photon energies is equal to the energy needed for the transition. The lack of use of the photosensitizer simplifies the procedures and prevents potential secondary effects, like excessive photosensitivity or unexpected phototoxicity. In a recent publication, we demonstrated the detection of phosphorescence at 1270 nm from the Raman photoexcited ¹O₂ [10]. However, determining the efficiency of the process (Φ_Δ) is still a task to be completed. The present work eliminates the gap. We define Φ_Δ as the relation between the number of Raman-generated ¹O₂ molecules and the total number of pumping photons. We measure Φ_Δ of (8±2) 10^-5 in distilled water (H₂O) when pumping at 410 nm with nanosecond pulses of an energy of 13 mJ.

Furthermore, we compare the Raman approach with the photosensitization method using the well-known photosensitizer, Rose Bengal. The ¹O₂ chemical trap, uric acid, is used to evaluate both methods. We show that, over time, the Raman method generates more ¹O₂ molecules than the photosensitization method. Rose Bengal exhibits near-full photobleaching after sixty minutes of irradiation, interrupting ¹O₂ photoproduction. In the Raman method, photobleaching does not take place. Consequently, the Raman ¹O₂ generation continues without interruption even after hours of irradiation.

2. Theoretical Considerations

Figure 1a shows a simplified schematic of ¹O₂ Raman excitation. A pumping photon induces a Raman transition from the ground toward the ¹O₂ level, emitting a Stokes photon. For example, when pumping at 410 nm in H₂O, the ¹O₂ Stokes component occurs around 605 nm. The magnitude of this signal yields a direct measurement of the amount of excited ¹O₂ molecules. Other Stokes photons related to the excitation of solvent vibrational modes and their overtones are generated. When pumping at 410 nm, the H₂O stretching vibrational mode produces a Stokes component at 475 nm. An additional overtone of this Stokes signal is also observed at 566 nm. The difference in wavelength between the Stokes components allows for their identification and separation using optical filters. Experiments below demonstrate that most of the energy involved in the Raman process originates from the solvent stretching modes. Despite this fact, the ¹O₂ Stokes can still be detected, showing that a significant amount of ¹O₂ molecules is still being generated.

In the photosensitization method, the photosensitizer’s quantum yield measures the efficiency of ¹O₂ photoproduction [11-15]. The photosensitizer’s quantum yield is defined as the number of excited oxygen molecules divided by the number of photons absorbed by the photosensitizer. The commonly used photosensitizer Rose Bengal exhibits a quantum yield value of 0.76 at 532 nm in H₂O [15-16]. As discussed above, for the Raman method, Φ_Δ is defined by the equation

$${\mathsf{\Phi }}_{\Delta }={\displaystyle \frac{N_{\Delta }}{N_p}},$$

(1)

where N_Δ and Np are the number of ¹O₂ Stokes and pumping photons, respectively. The number of photons can be estimated by measuring the signal energy corresponding to each process divided by the energy of one photon. Then, we can write

$${\mathsf{\Phi }}_{\Delta }={\displaystyle \frac{{\overline{\lambda }}_{\Delta }·E_{\Delta }}{{\lambda }_p\circ E_p}},$$

(2)

where ${\overline{\lambda }}_{\Delta }$ is the averaged ¹O₂ Stokes wavelength, E_Δ is its energy, λ_p is the pump wavelength, and E_p is its energy.

4. Results and Discussion

The solvent Stokes contributions are significant, but they can be eliminated using optical filters. Figure 2a shows the spectra collected using the Ocean Optics spectrometer after passing through neutral filters with a total optical density of six when pumping distilled water at 410 nm. The first peak at 410 nm corresponds to the pumping field. The peak at 475 nm corresponds to the H₂O stretching mode at a frequency of 3350 cm^-1 [20-23]. This mode is the dominant part of the total Stokes field.

Figure 2b shows the spectrum after using a neutral density filter with an optical density of two and a long-pass filter with a cutoff at 550 nm. A diminished Stokes peak at 475 nm and a peak at 566 nm are observed. The last peak corresponds to the first overtone of the H₂O vibrational mode at 6700 cm^-1. When using a neutral density filter with an optical density of one and a long-pass filter with a cutoff at 610 nm, the solvent Stokes components at 475 nm and 566 nm are fully depleted (see Figure 2c). The arbitrary units in all figures (2a, 2b, and 2c) are the same. Thus, the signal depicted by Figure 2c is four orders of magnitude smaller than the signal shown on Figure 2a and one order of magnitude smaller than the signal shown on Figure 2b. To demonstrate that the curve shown in Figure 2c is oxygen-dependent, we reduce the oxygen concentration using the oxygen quencher NaHSO₃.

Figure 3a shows the singlet peak obtained when using the long-pass filter at 610 nm for different concentrations of NaHSO₃. The blue line corresponds to the condition where no NaHSO₃ is added. As the NaHSO₃ concentration increases, the signal decreases. At a concentration of 40 mg/ml, the signal has been significantly reduced. Using the optical oxygen sensor, we measure the oxygen concentration in each sample solution. Figure 3b shows how the oxygen concentration decreases as the NaHSO₃ concentration increases. A nearly linear dependence is observed for concentrations below 10 mg/ml. Figure 3 shows that the observed Stokes signal collected using the 610 nm long-pass filter is oxygen-dependent. Its amplitude provides a direct measurement of the amount of ¹O₂ molecules being generated in the process. We note that the H₂O Stokes components are not affected by the presence of NaHSO₃.

Figure 4a shows the pulse energy corresponding to the excitation pump light (blue light). The signal was collected using the diode detector and energy sensor for calibration purposes. We use an interference filter at (410±10) nm to select only the pump light. Using an interference filter at (475±10) nm, we collect a Stokes signal corresponding to the H₂O stretching mode at 3350 cm^-1 (red line in figure 4a). A significant portion of the pump field, (23±1) %, is converted into this Stokes component. The standard deviation is estimated over 10 experiments. We also measured the contribution from the overtone at 566 nm using the corresponding filter, which accounted for (0.25±0.01) % of the total incoming light. The ¹O₂ Stokes contribution is significantly smaller. Figure 4b shows the ¹O₂ Stokes signal, together with the pumping signal, on a logarithmic scale. We estimated a pulse energy of (0.7±0.2) μJ. Under these conditions, equation (2) provides Φ_Δ=(8±2) 10^-5. Using these values, we estimate that each pulse of pumping light generates approximately 4 10^{11 1}O₂ molecules.

To understand the potential practical applications of the Raman method, we compare it with the conventional photosensitization method. We use the well-known photosensitizer Rose Bengal for this purpose. We prepare solutions of Rose Bengal in H₂O at a concentration of 10 μM. We also use uric acid at a concentration of 120 μM as a ¹O₂ chemical trap to monitor the amount of ¹O₂ being generated in each method. We irradiate the Rose Bengal and uric acid solution at 532 nm in five-minute intervals. At the end of each interval, we measure the UV spectrum of the solution to monitor the amplitude of the peak at 294 nm.

Figure 5a shows the UV-VI absorbance spectra of the Rose Bengal solution containing uric acid at a concentration of 120 μM, measured for different irradiation times. We observe a reduction in the uric acid peak at 294 nm, corresponding to the depletion of ¹O₂ generated by the uric acid. The peak depletes by a factor of four after twenty minutes but does not entirely disappear. The spectra also show the Rose Bengal absorbance peak around 540 nm. This peak also decreases, evidencing photobleaching. Due to the photobleaching of Rose Bengal, the process of ¹O₂ generation stops. Figure 5b shows results obtained for a solution of uric acid with no photosensitizer in H₂O. In this experiment, irradiation is performed at 410 nm over longer time intervals: 60, 90, 120, and up to 480 minutes. Again, we observe depletion of the uric acid peak at 294 nm, evidencing the Raman generation of ¹O₂ molecules. No photobleaching effect is observed in the Raman experiment, resulting in a more complete reduction of the 294 nm peak over time. Figure 5c shows the uric acid absorbance at 294 nm as a function of irradiation time for both experiments. For the Rose Bengal solution, the uric acid absorbance at 294 nm decreases from an initial value of 2.3 to 0.3 after 20 minutes of irradiation. It remains around this value for the rest of the experiment. For the Raman experiment, the peak reduces from 2.3 to a value around 0.1 after 400 minutes of irradiation. The Rose Bengal experiment exhibits a fast photoproduction of ¹O₂. However, due to photobleaching of the photosensitizer, the process is not fully complete. In the Raman experiment, the ¹O₂ photoproduction is less effective. However, because of the absence of photobleaching, the ¹O₂ photoexcitation persists as long as the sample is being irradiated.

5. Conclusions

We have estimated the efficiency of the ¹O₂ Raman photogeneration in H₂O by measuring the signal amplitude of the ¹O₂ Stokes signal. Defining the efficiency as the relation between the number of photogenerated ¹O₂ molecules divided by the number of pumping photons, we determine an efficiency of (8±2) 10^-4 for H₂O at 410 nm. We compare the Raman method with the photosensitization method using Rose Bengal as a photosensitizer. The photosensitizer generates ¹O₂ molecules rapidly, but the process eventually stops due to photobleaching of the photosensitizer. The Raman method generates ¹O₂ molecules at a rate slower than the photosensitization methods. However, thanks to the intrinsic absence of photobleaching, the Raman method generates more ¹O₂ molecules over time than the photosensitization method, despite its low efficiency values. The efficiency of the Raman method can be enhanced based on the fundamental principles of Raman interaction. Raman lasers, Raman amplifiers, and Raman fiber sensors are based on these principles [23-25]. Stokes signal grows exponentially with distance. The gain depends on the pumping field intensity, the nonlinear Raman susceptibility, and the propagation losses. A system should be designed to provide a significant gain for the ¹O₂ Stokes component with low losses, while maintaining low gain and substantial losses for the solvent Stokes components. These considerations may play a crucial role in improving the ¹O₂ Raman photogeneration efficiency for practical applications of the Raman approach. One useful application is the purification of water caused by viruses or bacteria. In a previous communication, we demonstrated the deactivation of viruses in an aqueous environment through simple irradiation with blue light, without the use of photosensitizers [26].