Properties of M(tpp).
Abstract
This chapter describes the photocatalysis action of (dihydroxo)tetraphenyl¬porphyrinato complexes of high valent P (V), Ge (IV), and Sb (V) (P(tpp), Ge(tpp), and Sb(tpp)). These chromophores were fixed onto silica gel (SiO2) through Coulombic forces and hydrogen bonding between axial hydroxo ligands and silanol groups to produce M(tpp)/SiO2 (M = P, Ge, and Sb) composites. M(tpp)/SiO2 were applied to the photo-inactivation of Escherichia coli and Legionella pneumophila. Moreover, M(tpp)/SiO2 was subjected to practical experiments for the photoinactivation of L. pneumophila naturally occurring in a cooling tower and a public fountain. It is noteworthy that 80 g of Sb(tpp)/SiO2 catalyst, containing 40 mg of Sb(tpp) maintained a concentration of Legionella species below 100 CFU/100 mL for 120 days in 13 m3 of water in a fountain under sunlight exposure. The photoinactivation proceeded through the liberation of M(tpp) from SiO2, adsorption of M(tpp) inside bacteria, and generation of reactive oxygen species, such as singlet oxygen, under visible light irradiation, thus resulting in bacteria apoptosis. Based on these results, we developed water-soluble porphyrins by modification of P and Sb porphyrin axial ligands to alkyloxo, alkylethylenedioxy, and alkylpyridinium groups. These water-soluble porphyrins were applied to the photodynamic inactivation of E. coli and Saccharomyces cerevisiae.
Keywords
- high valent metal
- P(V)-porphyrin
- Ge(IV)-porphyrin
- Sb(V)-porphyrin
- Escherichia coli
- Legionella pneumophila
- Saccharomyces cerevisiae
1. Introduction
Photocatalysis has received much attention as an environmentally friendly process for degrading organic compounds and bacteria in contaminated water. It is well known that UV-light irradiation of TiO2 generates a hydroxyl radical, which works as a strong oxidizing reagent for various microorganisms in aqueous solutions [1]. In 1985, Matsunaga et al. reported that Pt-loaded TiO2, under irradiation of >380 nm light, was capable of photoinactivation of various microorganisms, such as
In this chapter, we will cover M(tpp)/SiO2 and water-soluble porphyrins that were applied to the photosensitized inactivation of
2. Preparation of the photocatalysts
2.1. Porphyrin chromophores
The porphyrin chromophores studied were (dihydroxo) tetraphenylporphyrinatophosphorus chloride (P(tpp)), (dihydroxo)tetraphenylporphyrinatogermanium (Ge(tpp)), and (dihydroxo)tetraphenylporphyrinatoantimony bromide (Sb(tpp)). Axial hydroxo ligands were bonded to the metals through stable covalent bonds. P(tpp) and Sb(tpp) are cationic complexes, whereas Ge(tpp) is a neutral complex. Table 1 lists the physicochemical properties of the M(tpp)s, such as oxidation potentials (
M(tpp) | λmax/nmc ( | Fluorescence | Ref. | ||||
---|---|---|---|---|---|---|---|
Soret band | Q-band | λmax/( | |||||
P(tpp) | 1.20g | −0.93g | 424 (31.2) | 554 (1.82) | 607 (2.04) | 0.0416 (5.4) | [3] |
Ge(tpp) | 0.95 | −0.83 | 420 (77.6) | 554 (2.19) | 596 (2.08) | 0.1500 (4.7) | [4, 5] |
Sb(tpp) | 1.17 | −0.74 | 419 (41.6) | 552 (3.09) | 596 (2.08) | 0.0518 (1.6) | [6, 7] |
2.2. Preparation of porphyrins/silica gel composites
To perform the photoreaction in an aqueous solution, less water-soluble M(tpp) was fixed onto porous SiO2 to form M(tpp)/SiO2 [11]. Silica gel powder (300 mesh, 40 μm
2.3. Photocatalytic oxidation of organic compounds using M(tpp)/SiO2 (M = Ge, Sb)
Since M(tpp)/SiO2 (M = Ge, Sb) have high oxidation abilities, M(tpp)/SiO2 was applied to the oxidation of cycloalkenes [12] and acetone [4] and the dechlorination of 4-chlorophenol [13]. The photocatalytic reactions were performed using the setup depicted in Figure 3, where the reactant was supplied by a continuous flow system. In a spiral-type apparatus (Figure 3A), the reactant solution is fed continuously from a holder to a spiral glass tube packed with the photocatalyst. Sample irradiation was performed using a fluorescent lamp (22 W). The oval mirror-type apparatus (Figure 3B) consisted of a fluorescent lamp (18 W), an oval mirror, and a reactor (20 mmφ × 500 mm; 150 mL) packed with the photocatalyst. The fluorescent lamp was set on one focus of the oval mirror, and the reactor was set at another focus. The visible light emitted from the fluorescent lamp was concentrated onto the reactor. The reactant solution was fed continuously from reservoirs into the reactors.
The photocatalytic oxidations of cycloalkenes using oxygen were performed in a spiral type reactor (Figure 3A). Irradiation was directed onto a spiral glass tube (4 mm
Photo-oxidation of MeOH to HCHO was performed by Ge(tpp)/SiO2 at room temperature using a spiral type reactor (Figure 3A). Irradiation with a fluorescent light was performed on a spiral glass tube (4 mmφ × 2.5 m) packed with Ge(tpp)/SiO2 (12 g). The reactant was fed continuously into the spiral glass tube from the reservoir that contained an aerated aqueous solution of MeOH (50 mM in 200 mL) [4]. HCHO (30.6 μM) was formed after 180 h with a turnover number (TON) of 3.0 (Figure 5). The isotope effect for the photo-oxidation of methanol was found to be 2.1 from the ratios of slopes in the time-conversion plots for CH3OH and CD3OH. Therefore, it was suggested that the oxidation occurs through hydrogen abstraction. The generation of a Ge-O• species was generated by excitation of Ge(tpp). Here, Ge(tpp) acts as an O-radical generator. The photo-oxidation by Ge(tpp)/SiO2 was applied to toluene and ethylbenzene to produce alcohols and aldehydes/ketones. In the cases of cumene, methylcyclopentane, and methylcyclohexane, the corresponding alcohols were produced.
Dechlorination of 4-chlorophenol (4-CP) using Sb(tpp)/SiO2 was performed using the oval mirror-type apparatus (Figure 3B) [13]. Here, Fe(NO3)3 was used instead of O2 as the electron acceptor for the dechlorination of 4-CP, since the oxidation potential of 4-CP was relatively high. Sb(tpp)/SiO2 (0.087 wt% Sb(tpp)) was loaded into the oval mirror reactor. Before irradiation, the aqueous solution of 4-CP (initial concentration was 493 μM) was fed for 3 h under dark conditions, and the initial concentration of 4-CP decreased to 400 mM, probably due to the adsorption of 4-CP on the SiO2. Upon irradiation with the fluorescent lamp for 72 h, the concentration of 4-CP decreased from 400 to 6 μM along with the formation of Cl− (233 μM) and 1,4-benzoquinone (205 μM). Fe2+ (811 μM) was produced as a consequence of the reduction of Fe3+ (Figure 6). Electron transfer from the excited triplet state of Sb(tpp)/SiO2 to Fe3+ was responsible for the photodechlorination initiation, since Rehm-Weller equation calculated that the free energy change (ΔG) for the electron transfer from the excited triplet state of Sb(tpp) (
3. Photoinactivation of E. coli by M(tpp)/SiO2 (M = P, Sb)
Our first experiment for photoinactivation using Sb(tpp)/SiO2 was reported in 2003 against
Figure 8A shows the time courses of survival ratio (100
Similar photoinactivation studies of
4. Photoinactivation of Legionella species in naturally occurring environments
4.1. Photoinactivation of L. pneumophila
Initially, the photoinactivation of
Figure 8C shows the time courses for
4.2. Practical experiments in a cooling tower
The bactericidal experiment was performed in a cooling tower (Figure 9A) that was located in a building in Miyazaki city [18]. A cylindrical apparatus (200 mmφ × 500 mm, Figure 9B) that consisted of seven fluorescent lamps (18 W, 43 mmφ × 50 cm) and the Sb(tpp)/SiO2 catalyst (4.0 kg, 0.05 wt% Sb(tpp)) was used. Water in the holder (800 L) of the cooling tower was pumped into a cylindrical vessel at a rate of 28 L min−1, and then, the treated water was returned to the holder. The average retention time was calculated to be 26 s. In the cylindrical vessel, the Sb(tpp)/SiO2 catalyst was irradiated by visible light emitted from the fluorescent lamps at ambient temperature. Sampling of the water was carried out at outlet at 3–7 day intervals. At the same time, the atmospheric temperatures were recorded as the average value of the highest temperature of Miyazaki city during the sampling day and 2 days prior. The sample water (1.0 L) was filtrated through a membrane filter (0.45 μm, HA, Millipore) under reduced pressure into the vessel (100 mL) containing the microbes adhering to the membrane filter, an aqueous solution (5 mL) was added, and the vessel was shaken vigorously. Saturated aqueous KCl (5 mL, pH 2.2) containing 0.2 M HCl was added, and the vessel was shaken vigorously. After standing for exactly 20 min at room temperature, the prepared solution was ready for plating as described in Section 4.1. The amounts of
Under the conditions without any bactericidal treatments,
4.3. Practical experiments in water fountain
Practical experiments were performed in a public fountain of Miyazaki city (Figure 11A) that was filled with 13 m3 of water [18]. Photoinactivation of the fountain was examined using a leaf-type of photoinactivation apparatus (200 mmφ × 50 mm, Figure 11B) containing the Sb(tpp)/SiO2 catalyst (80 g, 0.05 wt% Sb(tpp)), which operated under sunlight irradiation. The determination of viable cell numbers of
As a result of the practical experiments, it is noteworthy that 80 g of Sb(tpp)/SiO2 catalyst, which contained 40 mg of Sb(tpp), could maintain the concentration of
4.4. Mechanism for photoinactivation using M(tpp)/SiO2
Elemental analyses of the catalysts before and after use in the fountain were performed with ICP. Before use, the Sb content in Sb(tpp)/SiO2 was measured to be 80 ppm, which was in good agreement with the Sb content (72 ppm) calculated for the 0.05 wt% of Sb(tpp) content in the catalyst. After 3 months of use in the fountain, the Sb content decreased from 80 to 17 ppm. On the other hand, Na, Mg, Al, and Ca largely increased, resulting in ion-absorption on SiO2. Moreover, Sb(tpp)/SiO2 catalyst used in the fountain was analyzed by a confocal laser scanning microscopy (CLSM). It was found that the fluorescence coming from the surface of the catalyst keep the similar shapes to the original catalyst, but the intensity was weaker compared with the original spectra of Sb(tpp)/SiO2. On the other hand, the fluorescence from the inside of the catalyst maintained the original intensity. Therefore, it is suggested that Sb(tpp) was eliminated from the surface of the catalyst. Irradiation of fluorescent light on the Sb(tpp)/SiO2 catalyst in deionized water did not sufficiently account for the spectral change and decrease in total Sb. Therefore, that the cationic Sb(tpp) chromophore was exchanged with alkali metal ions in the bulk water on the surface of the catalyst under irradiation is strongly suggested. A similar phenomena were observed in the case of P(tpp)/SiO2 [10]. Moreover, when Sb(tpp) was tightly fixed on SiO2 through covalent bonds, no photoinactivation occurred [19].
Therefore, the liberation of the Sb(tpp) chromophore from SiO2 is necessary for photoinactivation, as shown in Figure 13. Sb(tpp) can dissolve slightly in water (the water solubility (
5. Photosensitized inactivation using water-soluble porphyrins
As mentioned in the previous section, it was found that Sb(tpp) dissolved in water was responsible for the photoinactivation of bacteria. Therefore, in our next study, we intended to inactivate bacteria using water-soluble porphyrins. Water-soluble porphyrins have received much attention in connection with photoinactivation [20] and photodynamic therapy (PDT) [21, 22, 23, 24] ever since the first report on photoinactivation of
Entry | |||||||
---|---|---|---|---|---|---|---|
[ | [ | ||||||
0 | 0.10 | 50 | 380 | – | – | ||
1 | 0.13 | 50 | 192 | – | – | ||
6 | 1.09 | 50 | 14 | – | – | ||
10 | 2.10 | 40 | 22 | – | – | ||
12 | 2.21 | 50 | 17 | – | – | ||
14 | 2.40 | 50 | 21 | – | – | ||
1 | 3 | 17.4 | 500 | 23 | – | – | |
1 | 2 | 13.9 | 300 | 81 | – | – | |
2 | 2 | 13.0 | 200 | 31 | – | – | |
4 | 2 | 5.38 | 50 | 55 | – | – | |
6 | 2 | 2.07 | 5 | 64 | – | – | |
6 | 1 | 1.11 | 5 | 69 | – | – | |
2 | >120 | *50 | *32 | 250c | 32c | ||
4 | 112 | *50 | *36 | 250c | 53c | ||
6 | 63.6 | 50 | 44 | 250c | 120c | ||
1 | 3.35 | 50 | 20 | 2000 | 66 | ||
4 | 6.10 | *30 | *85 | 2000 | 27 | ||
5 | 3.80 | – | – | 500 | 29 | ||
6 | 5.84 | *20 | *72 | 500 | 31 | ||
7 | 6.00 | – | – | 400 | 24 | ||
8 | 3.80 | – | – | 500 | 63 |
We show the results of photoinactivation of
From the plots of the survival ratios against irradiation time, the bactericidal activity of porphyrins was evaluated by the half life (
6. Conclusion and perspectives
Since aqueous solutions are more transparent for visible light than ultraviolet, visible light photocatalysts work best for the photocatalytic reactions in aqueous solution. Moreover, visible light photocatalysts take advantage of the photocatalytic reactions under sunlight irradiation, since sunlight consists of 52% visible, 42% infrared, 6% UV-A, and 0.5% UV-B light. We showed two methods to photoinactivate bacteria: one method is the dispersion of M(tpp)/SiO2 (M = P, Ge, and Sb) in water, which is applicable to open system in naturally occurring environments; the other method is the water solubilization of M-porphyrins (M = P and Sb), which can be used in a closed system. M(tpp) and M-porphyrins can interact with bacteria through adsorption onto cell walls and absorption into the cells. Under irradiation, reactive species, such as 1O2, is generated by energy transfer from the porphyrins to O2 molecules on the cell walls and inside the cells.
Thus, porphyrins are useful chromophores for catalysis and sensitization in a biological application. The application of water-soluble M-porphyrins to PDT is currently underway in our laboratories.
Acknowledgments
This work was partially supported by a Grant-in-Aid for Scientific Research (C) (16K05847) from Japan Society for the Promotion of Science (JSPS). Also, the authors would like to express grateful acknowledgment to many students belonged in our laboratory during 2003 to present when visible light inactivation has been investigated in our laboratory for their large contributions.
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