MECHANISM OF THE PHOTOCHEMICAL REACTION OF DIPHENYLAMINES WITH CARBON TETRACHLORIDE

Abstract. The quantum yields of HCI (φ_HC1) formation have been measured for the photolysis of N -methyldiphenylamine (MeDPA), triphenylamine (TPA) and diphenylamine (DPA) in the presence of CCl₄ in polar solvents. The quantum yields of N-methylcarbazole formation (φ_mφca) have also been determined for the system MeDPA-CCl₄. With increasing CCl₄ concentration, φ_HCl increases as φ_MeCA decreases, and φ_HCl reaches maximum values 2.7 at 1 M CCl₄. Using laser photolysis, transient spectra have been recorded for MeDPA in the absence and presence of CCl₄ in polar and non-polar solvents, and for TPA. Transient absorption due to the triplet states and photocyclization products (without CCU), exciplexes, the (C₆H₅)₂ NCHi radical, the MeDPA⁺ cation radical, the (TPA⁺., CCl₄) ion pair, and the TPA⁺ cation radical have been identified. The mechanistic implications of these results are discussed.     

PHOTOLYSIS OF SUBSTITUTED NAPHTHALENES ON SiO2 AND Al2O3

Abstract— Photolysis of naphthalene on the surface of SiO₂ under an atmosphere of air produces phthalic acid as the only major photoproduct, accounting for 49%o of the consumed naphthalene. Photolysis on Al₂O₃ also produces phthalic acid, in 31% yield. Photolysis of 1 -methylnaphthalene on SiO₂ proceeds under similar conditions to produce 2-acetylbenzoic acid (35%) as the major photoproduct with the production of a small amount of I-naphthaldchyde (6%). 1-Cyanonaphthalene does not photooxidize under similar conditions. The presence of oxygen is necessary for the photodecomposition of naphthalene and 1-methylnaphthalene to proceed. Superoxide formed from the photolysis of naphthalene at the SiO₂/air interface is readily observed by electron paramagnetic resonance spectroscopy. In the absence of naphthalene no superoxide is observed. A mechanism involving electron transfer from the S₁ state of the naphthalene to O₂ is proposed on the basis of these observations and related literature precedent.     

LUMIFLAVIN-SENSITIZED PHOTOOXYGENATION OF INDOLE

Abstract— The lumiflavin-sensitized photooxygenation of indole in aqueous solutions has been investigated by means of steady light photolysis and flash photolysis. The semiquinone of lumiflavin and the half-oxidized radical of indole were formed by the reaction between triplet lumiflavin and indole (3.7 times 10⁹ M ^-1 s^-1). The semiquinone anion radical of lumiflavin reacted with oxygen to form superoxide radical. The triplet state of lumiflavin also reacted with oxygen forming singlet oxygen, ¹O₂. But the reaction between ¹O₂ and indole (7 times 10⁷ M^_l s^_1; estimated from steady light photolysis using Rose Bengal as a sensitizer) was far less efficient than the reaction between indole and triplet lumiflavin. The quantum yield of the lumiflavin-sensitized photooxygenation of dilute indole via radical processes was much higher than that via ¹O₂ processes, though appreciable ¹O₂ was formed.     

RESONANCE RAMAN and ABSORPTION SPECTRA OF ISOMERIC RETINALS IN THEIR LOWEST EXCITED TRIPLET STATES

Abstract— The triplet-triplet absorption spectra of 9-cis-, 13-cis- and all-fraw-retinal as well as the time-resolved resonance Raman spectra of the lowest electronically excited triplet states of 9-cis-, 11-cis, 13-ciy and all-trans-retinal in aromatic solvents at room temperature have been obtained under conditions ensuring the isomeric purity of the starting materials. The triplet states were produced by triplet energy transfer from a sensitizer in pulse-radiolysis experiments. The overall results suggest that the isomeric retinals form either different relaxed triplet species or different mixtures of relaxed triplet species. The possible implications about the size of the energy barriers separating the various triplet species are discussed. The resonance Raman spectra obtained by using either anthracene (E_T= 177.7 kJ mol^-1) or naphthalene (E_T= 254.8 kJ mol^-1) as sensitizers were virtually identical for the corresponding triplet states from each of the isomers 11-cis-, 13-a's- and all-tams-retinal, suggesting that the relaxed triplet species or the mixture of relaxed triplet species formed from each isomer is independent of the energy of the sensitizer.     

EFFECT OF AGGREGATION ON THE HEMATOPORPHYRIN-SENSITIZED PRODUCTION OF SINGLET MOLECULAR OXYGEN

Abstract— The hematoporphyrin-sensitized production of singlet molecular oxygen, O₂(¹Δ_g), has been investigated in methanol and in aqueous solution. The quantum yield for formation of O₂(¹Δ_g) (Φ_Δ) has been measured by both steady-state (oxygen consumption) and time-resolved (near-infrared luminescence) methods. In methanol, both techniques indicate that Φ_Δ= 0.76 and the value remains independent of sensitizer concentration over a wide range. This finding is consistent with the dye persisting in a monomelic form in methanol solution. In contrast, Φ_Δ decreases markedly with increasing sensitizer concentration in water due to dimerization of the dye. Analysis of the steady-state data indicates Φ_Δ values of 0.74 and 0.12, respectively, for monomer and dimer. It is further shown that the efficiency whereby quenching of the triplet state by O₂ results in generation of O₂(¹Δ_g) is substantially lower for the dimer than for the corresponding monomer. Because monomer and dimer possess quite different absorption spectral profiles, the efficacy for photodynamic action with hematoporphyrin exhibits a pronounced wavelength dependence.     

PHOTODYNAMIC INACTIVATION OF E. COLI BY ROSE BENGAL IMMOBILIZED ON POLYSTYRENE BEADS

Abstract. The photodynamic inactivation of E. coli by visible light and O₂ was found to occur in the presence of the sensitizer rose bengal, immobilized by covalent bonding to polystyrene beads. The demonstrated absence of significant amounts of dissolved rose bengal indicated that an inactivation mechanism based on penetration of sensitizer molecules into the cell's interior could not be operating. Survival curves typically exhibited induction periods followed by rapid exponential death, with 99.99% kill requiring 1–2 h depending on conditions. A mechanism involving the participation of photo-generated singlet excited oxygen O₂(¹δ) in inactivation of E. coli is proposed. The photodynamic inactivation rate increased significantly in H₂O compared with H₂O, which is evidence supporting singlet oxygen as an active intermediate, since O₂(¹δ) has a much longer lifetime in H₂O than in H₂O. H₂O did not act as a short term poison in the absence of sensitizer.     

PHOTOBLEACHING OF MONO-l-ASPARTYL CHLORIN e6 (NPe6): A CANDIDATE SENSITIZER FOR THE PHOTODYNAMIC THERAPY OF TUMORS

Abstract— Most sensitizers used for the photodynamic therapy (PDT) of tumors photobleach on illumination. Thus, it is of interest to examine the photobleaching behavior of new sensitizers proposed for use in PDT. This report surveys the quantum yields and kinetics of the photobleaching of mono- l -aspartyl chlorin e₆ (NPe6), a hydrophilic chlorin that has many of the photoproperties desirable in a sensitizer for clinical PDT. It is a very effective sensitizer for the PDT of several types of model tumors in animals and is now in Phase I clinical trials. The quantum yield of NPe6 photobleaching in pH 7.4 phosphate buffer in air was 8.2 × 10⁻⁴; this is greater than the yields for typical porphyrin photosensitizers. For example, the yields for hematoporphyrin and uroporphyrin are 4.7 × 10 ⁵ and 2.8 × 10⁻⁵, respectively. The yield decreased significantly in organic solvents of low dielectric constant. The Sn derivative of NPe6 was more light stable than NPe6 (yield = 5.7 × 10 ⁻⁶), while the Zn derivative was more sensitive (yield = 1.9 × 10⁻²). Oxygen appeared to be necessary for the photobleaching of NPe6; however, bleaching was not inhibited by 100 mM azide, an efficient quencher of singlet oxygen. The photooxidizable substrates cysteine, dithiothreitol and furfuryl alcohol increased the quantum yield of photoblcaching two- to four-fold, while the electron acceptor, met-ronidazole, increased it almost six-fold. Photobleaching yields for several other chlorins were also measured.     

SENSITIZED PHOTOOXYGENATION ACCORDING TO TYPE I MECHANISM (RADICAL MECHANISM) — PART III. EXPERIMENTS WITH CONTINUOUS ILLUMINATION

Abstract— The photochemical reaction in the system thionine (sensitizer), allylthiourea (ATU, acceptor), and oxygen was studied with continuous illumination. In oxygen-free aqueous solution thionine is photoreduced to leucothionine. With oxygen, however, a photooxygenation of the acceptor takes place. At the same time the quantum yield of the bleaching reaction of thionine decreases markedly in comparison with that of the oxygen-free solution. At about 100 sec after the beginning of illumination, the overall quantum yield of the bleaching reaction diminishes further because the leucothionine formed during the reaction now becomes transformed into thionine. The quantum yields do not change significantly over the range of oxygen concentrations studied (initial concentration 1 × 10^-5 to 5 × 10^-5 M ). In addition they are independent of the light intensity. The influence of the pH and the acceptor concentration were also investigated.
The sensitizer is not only bleached reductively, but is also partly destroyed by oxidation. The results are in agreement with the reaction scheme elucidated by flash photolysis measurements.
In accordance with this reaction scheme, the primary reaction (a) of the reductive bleaching of the sensitizer, (b) of the photooxygenation of the acceptor and (c) of the oxidative destruction of the sensitizer, is identical in all cases. This process is the redox reaction between the sensitizer triplet and the acceptor, where a semithionine and an ATU-radical are formed. The reaction represents an example of a Type I photooxygenation according to the notation of Gollnick.     

POLYMER BOUND PYRROLE COMPOUNDS–VI. PHOTOPHYSICAL PROPERTIES OF MONOMERIC MODELS FOR POLYSTYRENE-BOUND PORPHYRINS

Abstract— Several porphyrin esters used as models for polystyrene-bound porphyrins have been prepared and their excited states have been studied by laser flash photolysis, IR phosphorescence of singlet molecular oxygen, O₂(¹Δ_g), and steady-state fluorescence. The photophysical properties of the porphyrin esters in solution are affected by the presence of nitro group(s) in the chain. In this case, an important decrease in φ_f, φ_T and φ_δ (to ca 0.7–0.4 of the value for the parent dimethyl ester) is observed. This is mainly due to intramolecular electron-transfer quenching [by the nitro group(s)] of the first excited singlet state of the porphyrin. The thermodynamic feasibility of this deactivation pathway has been confirmed polarographically. Quenching of the porphyrin triplet state and of O₂(¹Δ_g) by the nitro groups is negligible. The present conclusions explain also the results obtained previously for the photooxidation of bilirubin sensitized by the parent insoluble polystyrene-bound porphyrins. In that case the photooxidation rates were correlated directly with the quantum yield of O₂(¹Δ_g) production by the sensitizer. The consequences of these results for the use of polystyrene-bound porphyrins in sensitized photooxidation processes are discussed.     

PHOTODYNAMIC INACTIVATION OF LYSOZYME BY EOSIN

Abstract— It has been demonstrated that singlet oxygen is the major oxidizing entity in the photo-dynamic inactivation of hen egg white lysozyme by eosin, using D₂O to enhance the solvent-induced decay lifetime, and azide ion as a specific scavenger. Two regimes of inactivation can be distinguished depending on whether the sensitizer is free or complexed to the enzyme. The kinetic analysis for free dye sensitization, based on photostationary measurements and inactivation quantum yields, indicates that at least 1 in 15 singlet oxygen interactions with lysozyme leads to loss of lytic activity. The direct attack of triplet eosin makes a lesser overall contribution in air-saturated solutions, where 1 in 4 reactions induces inactivation. Lysozyme binds 1 eosin molecule from pH 4 to 12, leading to almost total quenching of the tryptophyl residue fluorescence without inhibition of the enzymic activity. The inactivation quantum yields indicate that singlet oxygen generated from the bound dye is the inactivating agent, but the dominant attack takes place with the complexed fraction of lysozyme molecules. The tryptophyl residue loss is the same or smaller in changing from H₂O to D₂O despite the 5–10 times increase in quantum yield, indicating that singlet oxygen inactivates also by reacting with residues other than tryptophan. The photochemical and fluorescence results are consistent with the the identification of tryptophyl site 108 with the eosin binding site and a reaction target for singlet oxygen. In a re-examination of earlier work on eosin-sensitized photo-oxidation of I", it has been found that singlet oxygen is the oxidizing agent in aerobic solutions.     

EXCITED STATES OF BILIRUBIN

Abstract— The excited states of bilirubin (BR) in a variety of environments have been studied by 347 nm laser flash photolysis. Quantum yields of formation of triplet BR have been shown to be less than 0.005 in solution in water ( p H 9–11), methanolic ammonia, 10% aqueous mulgofen and in cetyl trimethyl-ammonium bromide. In benzene the quantum yield was 0.01 although this diminished to less than 0.005 on addition of triethylamine. Permanent products are formed with benzene and with 1% methanolic ammonia. With BR in HSA a transient decaying with k = 3.5 × 10⁵ s^-1 is formed by a monophotonic process together with a permanent product. Neither species is affected by oxygen or by iodide ion. Both originate from BR molecules in the strongest binding site in the HSA. The yields of both species are unaffected by salt but are temperature dependent. The decay of the transient is strongly temperature dependent corresponding to an activation energy of about 50–60 kj mol^-1. If this transient is a triplet it is formed with a quantum yield of 0.13 ± 0.01. The relevance of these results to an understanding of the photo therapeutic process is discussed.     

Comparative Photophysical Study of Disulfonated Aluminum Phthalocyanine in Unilamellar Vesicles and Leukemic K562 Cells

Abstract— The photophysical properties of cis -disulfonated aluminum phthalocyanine (AlPcS₂) in unilamellar vesicles (liposomes) of DL-a-dipalmitoyl-phosphatidylcholine have been measured. Both the fluorescence and triplet quantum yields decreased with increasing sensitizer concentration. The time-resolved fluorescence decays, analyzed by both the sum of exponentials and decay time distribution analyses, are compared with those reported for AlPcS₂ in leukemic K562 cells. Information on the pho-todynamic transport and localization mechanism has been obtained by drawing correlations between the two systems, indicating active transport of the phthalocyanine into tumor cells involving lysosomal accumulation.     

MECHANISMS OF THE PHOTOSENSITIZED OXIDATION OF TYRAMINE

Abstract— …According to the criteria of enhancement in D₂O and inhibition by sodium azide, the oxidation of tyramine photosensitized by methylene blue is largely a singlet oxygen or Type II process. Its quantum yield approximates 0.3 in D₂O at pH 10. There is a less efficient reaction not quenched by azide, which is assigned to a dye-substrate or Type I process. It gives rise to products with distinct bands at 320 and 285nm. Products of the Type I reaction are further oxidized by singlet oxygen and thereby compete with tyramine for this reagent. Kinetic parameters were estimated by computer simulation of the dependence of quantum yield on extent of reaction. The rate constant for reaction of O₂ (¹Δ_g) with tyramine was estimated to be 2.8 × 10⁸ M ^-1 s ^-1± 20% at pH 10. The reaction was also sensitized by hypericin in what appears to be a Type II process.     

FORMATION AND CHEMILUMINESCENT DECOMPOSITION OF DIOXETANES IN THE GAS PHASE

Abstract— High resolution chemiluminescence spectra have been obtained of the singlet electronically excited products of O₂(¹Δ) plus alkene, dioxetane forming, reactions. The experiments were conducted in a flow apparatus at pressures of 1–5 torr. The spectra are a measure of the unrelaxed initial distribution of energy in the excited product. Results are reported for ethylene, 1, 1-difluoroethylene. methyl vinyl ether, ethyl vinyl ether, n -butyl vinyl ether, ketene, ketene-d₂, allene, unsymdimethyl allene, dimethyl ketene, 2-methoxy propene, 1-ethoxy propene, 2-bromo propene, and N, N- dimethyl isobutenyl amine. Chemiluminescence activation energies, representing the cycloaddition process, and absolute quantum yields for singlet excited product, ranging from 10^--⁴ to 2.5 × 10^--². are reported for 10 alkenes. Several of the reactions, 1,1-difluoroethylene, ketene, ethylene and allene give formaldehyde ¹ nπ* product with excess vibrational-rotational energy and a higher quantum yield than reactions not displaying this phenomenon. This is an indication of at least partially statistical partitioning of the energy in excess of that needed to electronically excite the formaldehyde. The experiments with ketene and ketene-d₂ provide the first evidence for the existence of unsubstituted 1,2-dioxetanone. The results from several of the experiments, particularly those with 2-methoxy propene and I-ethoxy propene are consistent with the mechanism of Goddard, which predicts regioselective and stereoselective attack of O₂(¹Δ) upon alkoxy substituted alkenes having allylic hydrogen.     

USE OF TRIS (2,2'-BIPYRIDINE) RUTHENIUM(II) DICATION IN A NOVEL METHOD FOR THE DETERMINATION OF SINGLET OXYGEN REACTION RATES: APPLICATION TO STUDIES OF GELATIN

Abstract— A novel method for the determination of singlet oxygen reaction rate constants is described and applied to studies of cyclohexadiene in methanol and gelatins in H₂O and D₂O. The technique uses tris (2,2'-bipyridine) ruthenium(II) dication (Ru(bipy)₃₂₊) as both singlet oxygen sensitizer and in situ oxygen concentration monitor during irradiation of sealed samples. Because of the high efficiency with which the luminescence of Ru(bipy)₃2+* can be detected, and the fact that emission lifetimes are used, the method offers some advantages over those previously described. The advantages and disadvantages of the method are discussed. A rate constant of 2.1 (±0.3) x 10⁶ mol^-1 dm³ s^-1 has been determined for the reaction of ¹O₂ with cyclohexadiene in methanol. For two different photographic gelatins the sums of reaction and quenching rate constants are 2.0 (±0.4) x 10⁶ and 3.1 (±2.0) x 10⁵ mol^-1 dm³ s^-1; for swine skin gelatin this value is 3.9 (±2.4) × 10⁵ mol^-1 dm³ s^-1. Chemical reaction, rather than physical quenching, is the dominant reaction route for gelatins and ¹O₂.     

FLUORESCENCE OF 5-METHYLCYTOSINE

Abstract— 5-Methylcytosine and 5-methyldeoxycytidylic acid are fluorescent in aqueous solution at room temperature and neutral pH. 5-Methylcytosine, 10^-3M, pH 8.5, 25°C, has a quantum yield of 5 ×10^-4, 5-Methyldeoxycitydylic acid, 10^-4M, pH 7.5, 20°C, has a quantum yield of 8 × 10^-4. Emission maxima are 2.91 and 2.80μ^-1. At pH 14, the quantum yield of 5-methylcytosine is 1.6 × 10^-2; the emission maximum is 2.82μ^-1. At pH I, the quantum yield of both compounds is less than or equal to 10^-4. Both compounds were chromatographically homogeneous, had absorption spectra which agreed with published data, and excitation spectra which agreed closely with absorption spectra.     

THE SUSCEPTIBILITY OF MUNG BEAN CHLOROPLASTS TO PHOTOINHIBITION IS INCREASED BY AN EXCESS SUPPLY OF IRON TO PLANTS: A PHOTOBIOLOGICAL ASPECT OF IRON TOXICITY IN PLANT LEAVES

In an attempt to elucidate the underlying mechanisms for iron toxicity in plants, the combined effects of iron overload and light intensities on the photosynthetic capacity of leaves were particularly focussed upon in this study, using mung bean seedlings grown under varied conditions regarding the supply of light and iron. The seedlings, when supplied with excess iron (up to 1.0 m M ) and low light (40 W/m²), did not suffer any loss of photosynthesis; further, the typical symptoms of iron toxicity, as shown in the leaves grown in sunlight at ca 450 W/m² on an average, were not seen in those. Nonetheless, excess iron supply resulted in a marked increase in photosensitivity of the low light-adapted seedlings. A large portion of iron accumulated in chloroplasts by the supply of excess iron was found to be incorporated into thylakoids as nonheme iron (NHI), which acts as a potent sensitizer, photogenerating singlet oxygen (¹O₂). The generation rate of ¹O₂ from thylakoids linearly increased with increasing content of NHI; this was in parallel with the NHI content dependence of photoinactivation rates of photosynthetic electron transport and key enzymes of the Calvin cycle in chloroplasts. The results suggest that Fe-dependent photosensitization reactions, occurring via the ¹O₂ mechanism, may be deeply involved in cellular processes leading to developing iron toxicity symptoms in plants.     

Photochemical Reactions Involved in the Phototoxicity of the Anticonvulsant and Antidepressant Drug Lamotrigine (Lamictal®)

Lamotrigine (LTG) [3,5-diamino-6-(2,3-dichlorophenyl)-1,2,4-triazine], an anticonvulsant and antidepressant drug Lamictal^®, produces a (photo)toxic response in some patients. LTG absorbs UV light, generating singlet oxygen (¹O₂) with a quantum yield of 0.22 in CH₂Cl₂, 0.11 in MeCN and 0.01 in D₂O. A small production of superoxide radical anion was also detected in acetonitrile. Thus, LTG is a moderate photosensitizer producing phototoxicity and oxidizing linoleic acid. LTG is a weak ¹O₂ quencher ( k _q = 3.2 × 10⁵  m ⁻¹ s⁻¹ in MeCN), but its photodecomposition products in dimethyl sulfoxide (DMSO) quenched ¹O₂ very efficiently. Upon intense UV irradiation from a xenon lamp, LTG was photobleached rapidly in DMSO and slowly in acetonitrile, alcohol and water. The rate increased significantly when laser pulses at 266 nm were employed. The photobleaching products generated ¹O₂ twice as strongly as LTG. Photobleaching was usually accompanied by the release of chloride anions, which increased in the presence of ascorbic acid. This suggests the formation of aryl radicals via dechlorination, a process which may be responsible for the photoallergic response observed in some patients. Our results demonstrate that LTG is a moderate generator of ¹O₂ prone to photodechlorination, especially in a reducing environment, which can contribute to the reported phototoxicity of LTG.     

pH DEPENDENCE OF SINGLET OXYGEN PRODUCTION IN AQUEOUS SOLUTIONS USING THIAZINE DYES AS PHOTOSENSITIZERS

Abstract— The production of singlet oxygen by thiazine dye photosensitization, as measured by the rate of photooxidation of tryptophan, was found to be very sensitive to changes of pH in the range 5–9. For methylene blue in aerated solutions, the production of ¹O₂* is approximately five times more efficient in basic than in acidic medium. This was shown to be related to the p K 's of the triplet dyes, by evaluating the yields of ¹O₂* from the lifetimes and the quenching rate constants for the two ionic species of sensitizer triplets measured by laser flash photolysis. Changes in the quenching rate constants of the thiazine triplet states can be correlated with the triplet energies.