Reactive Oxygen Species Generation Sensitized By Riboflavin:Mechanism And Applications

The generation of reactive oxygen species (ROS) is a key technology in wastewater treatment. Simple, effective and low-cost methods for ROS generation are being pursued. This work was designed to develop a novel, precisely-controllable ROS generation method using the photosensitization of riboflavin (RF) under visible irradiation. In this dissertation, the photosensitization process of RF and the mechanism of ROS generation were investigated using electron paramagnetic resonance (EPR) combined with spin trapping; high performance liquid chromatography (HPLC), high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Since RF is an important widely-present natural sensitizer, an elucidation of RF-initiated ROS generation mechanism would also help us understand the aqueous photochemical behavior of bio-refractory organic pollutants in natural environments. Main contents and results are as follows:1. The generation of superoxide radical, sensitized by RF under visible irradiation, was validated by EPR combined with spin trapping. In order to further validate the generation of superoxide radical, photodegradation experiments of methyl orange (MO) were carried out in the presence of singlet oxygen quencher (1,4-diazabicyclo[2.2.2]octane) and superoxide anion radical quencher (1,4-benzoquinone). Only 1,4-benzoquinone was able to inhibit the photodegradation, suggesting that the photooxidation of MO was mainly initiated by superoxide radical.2. A highly efficient visible-light-mediated hydroxyl radical (HO·) generation method using the photosensitization of RF catalyzed by ferrous ion was developed. The generation of HO· was validated by EPR using 5,5-Dimethyl-l-Pyrroline-N-Oxide (DMPO) as spin trapping agent, and was further confirmed by using the hydroxylation of nitrobenzene as a model reaction. All reagents involved are nontoxic, environmentally benign, and biocompatible. Thus, this approach has a broad application.3. RF is not photo-stable and can be rapidly photodecomposed under both ultraviolet and visible irradiations. The photoproducts of RF under different pHs were analyzed using HPLC and HPLC-MS/MS. Experimental results clearly show that RF was degraded completely in one hour under solar irradiation, with LF and LC as its two main photoproducts.4. It is usually presumed that the most abundant HO* generation in-vivo is the transition-metal-ion catalyzed Fenton decomposition of hydrogen peroxide (H2O2). EPR spectroscopy was used to examine the effects of catalase on the HO· generation. The signal intensities of DMPO/HO· decreased substantially in the presence of catalase. With the analysis above, a possible mechanism for RF-mediated HO· generation catalyzed by iron-ions under visible irradiation is proposed. First, superoxide radical is produced under visible irradiation in the presence of RF. However, superoxide radical is sensitive in the presence of proton and is converted to H2O2 quickly in water. Ultimately, the HO· generation occurs through iron-ion catalyzed decomposition of H2O2. The experimental results clearly demonstrate that HO· can be generated under visible irradiation in the presence of an endogenous sensitizer (e.g., RF in this case) and iron ion (Fe2+).5. The ROS generation sensitized by other flavins, e.g., flavin adenine dinucleotide, FAD; Flavin mononucleotide, FMN; lumiflavin, LF; lumichrome, LC, was investigated using EPR combined with spin trapping. It was demonstrated that only LC could not initiate ROS generation under visible irradiation. To find the possible reason, UV-visible spectroscopy and three-dimensional excitation-emission matrix fluorescence spectroscopy were applied to analyze the structures of flavins, and an obvious blue-shift was observed in spectra of LC.6. The ROS generation method has been used to treat several typical bio-refractory organic pollutants (methyl orange; nitrobenzene; diclofenac); and to simulate the photosensitization of RF in natural environment. The elucidation of RF-initiated ROS generation mechanism will also help us understand the photochemical behaviors of organic pollutants and the biogeochemical cycling of elements in natural environments.