Formation Of Singlet Molecular Oxygen Research Articles

Photosensitized Formation of Soluble Bionanoparticles of Lysozyme

AbstractPhotosensitized crosslinking of hen egg‐white lysozyme (LZ) in buffer solutions was obtained by blue‐LED excitation of ruthenium (II) tris‐bipyridyl dication (Ru(bpy)32+) in a molar excess of persulfate anion (S2O82−), both in the absence and presence of dissolved oxygen. The results show a rapid monomer consumption of >75 %, with sequential formation of soluble LZ nanoparticles, e. g., dimers (LZ2),…n‐mers (LZn≥6) with diameters from 6 to 13 nm, regardless of the presence of dissolved oxygen in solution. The bionanoparticles are crosslinked through 3,3'‐dityrosine (diTyr) bonds, by the recombination of tyrosyl radicals of Y20, Y23, and Y53 residues of different protein molecules, produced by intermediated oxidants Ru(bpy)33+ and SO4.− generated after excitation. Furthermore, under aerobic conditions, photosensitized formation of singlet molecular oxygen (1O2) also results in the oxidative transformation of tryptophan residues into fluorescent kynurenine‐type derivatives. These findings may contribute to the rational development of functional protein bionanoparticles.

Excited States of Thio-2′-deoxyuridine Bearing an Extended π-Conjugated System: 3′,5′-Di-O-acetyl-5-phenylethynyl-4-thio-2′-deoxyuridine

A new thio-2'-deoxyuridine with an extended π-conjugated group was successfully synthesized: 3',5'-di-O-acetyl-5-phenylethynyl-4-thio-2'-deoxyuridine. The thio-2'-deoxyuridine derivative has a large red-shifted absorption band in the UVA region and also shows fluorescence, a rare photo-property among thionucleobases/thionucleosides. The triplet-triplet absorption spectrum and the rate constants (the intrinsic decay rate constant of the triplet state, the self-quenching rate constant, and the quenching rate constant of the triplet state by an oxygen molecule) of the thio-2'-deoxyuridine were obtained by transient absorption spectroscopy. The quantum yield of intersystem crossing and the quantum yield of singlet molecular oxygen formation (ϕΔ) under an oxygen atmosphere were also determined. The ϕΔ value of the new thio-2'-deoxyuridine was found to be substantially higher than all reported values of other thio-2'-deoxyribonucleosides in low oxygen concentrations similar to cancer cell environments. The fluorescence quantum yield depended on the excitation wavelength, revealing certain photochemical reactions in the higher excited singlet states. However, when excited into the higher excited state with non-resonant two-photon absorption, the ϕΔ of the thio-2'-deoxyuridine derivative was found to remain sufficiently large. These findings should be very useful for the development of thio-2'-deoxyribonucleoside-based pharmaceuticals as DNA-specific photosensitizers for photochemotherapy.

Magnetic Nanoplatforms for in Situ Modification of Macromolecules: Synthesis, Characterization, and Photoinactivating Power of Cationic Nanoiman–Porphyrin Conjugates

A nanoplatform concept was developed to synthesize accessible photoactive magnetic nanoparticles (MNPs) of Fe3O4 coated with silica. This approach was based on the covalent binding of 5,10,15,20-tetrakis(pentafluorophenyl)porphyrin (TPPF20) to aminopropyl-grafted MNPs by nucleophilic aromatic substitution reaction (SNAr) to obtain conjugate MNP-P1. After in situ modification, the remaining pentafluorophenyl groups of TPPF20 attached to MNPs were substituted by dimethylaminoethoxy groups to form MNP-P2. The basic amine group of these conjugates can be protonated in aqueous media. In addition, MNP-P1 and MNP-P2 were intrinsically charged to produce cationic conjugates MNP+-P1 and MNP+-P2+ by methylation. All of them were easily purified by magnetic decantation in high yields. The average size of the MNPs was ∼15 nm, and the main difference between these conjugates was the greater coating with positive charges of MNP+-P2+, as shown by the zeta potential values. Absorption spectra exhibited the Soret and Q bands characteristic of TPPF20 linked to MNPs. Furthermore, these conjugates showed red fluorescence emission of porphyrin with quantum yields of 0.011-0.036. The photodynamic effect sensitized by the conjugates indicated the efficient formation of singlet molecular oxygen in different media, reaching quantum yield values of 0.17-0.34 in N,N-dimethylformamide. The photodynamic activity of the conjugates was evaluated to inactivate the Gram-positive bacteria Staphylococcus aureus, the Gram-negative bacteria Escherichia coli, and the yeast Candida albicans. The modified cationic MNP+-P2+ was the most effective conjugate for photodynamic inactivation (PDI) of microorganisms. Binding of this conjugate to bacteria and photoinactivation capability was checked by means of fluorescence microscopy. Also, sustainable use by recycling was determined after three PDI treatments. Therefore, this methodology is a suitable scaffold for the in situ modification of conjugates, and in particular, MNP+-P2+ represents a useful photodynamic active material to eradicate microorganisms.

Polyethylene glycol-coated blue-emitting silicon dots with improved properties for uses in aqueous and biological environments

Grafting of polyethylene glycol (PEG) to ultrasmall photoluminescent silicon dots (SiDs) is expected to improve and expand the applications of these particles to aqueous environments and biological systems. Herein we report a novel one-pot synthesis of robust, highly water compatible PEG-coated SiDs (denoted as PEG-SiDs) of (3.3 ± 0.5) nm size. The nanoparticles’ synthesis is based on the liquid phase oxidation of magnesium silicide using PEG as reaction media and leading to high PEG density grafting. PEG-SiDs enhanced photophysical, photosensitising, and solution properties in aqueous environments are described and compared to those of 2 nm size PEG-coated SiDs with low PEG density grafting (denoted as PEG-NHSiDs) obtained from a multistep synthesis strategy. PEG-SiDs form highly dispersed suspensions in water showing stable photoluminescence and quantum yields of Φ = 0.13 ± 0.04 at 370 nm excitation in air-saturated suspensions. These particles exhibited the capacity of photosensitising the formation of singlet molecular oxygen, not observed for PEG-NHSiDs. PEG robust shielding of the silicon core luminescent properties is further demonstrated in bio-imaging experiments stressing the strong interaction between PEG-SiDs and Staphylococcus aureus smears by observing the photoluminescence of particles. PEG-SiDs were found to be nontoxic to S. aureus cells at concentrations of 100 mg ml−1, though a bacteriostatic effect on S. aureus biofilms was observed upon UV-A irradiation under conditions where light alone has no effect.

Photoinduced Energy and Electron-Transfer Reactions by Polypyridine Ruthenium(II) Complexes Containing a Derivatized Perylene Diimide

The [Ru(II) (phen)2(pPDIp)]2+ complex, where pPDIp is the symmetric bridging ligand phenanthroline–perylene–phenanthroline, shows strong electronic absorption bands attributed to the pPDIp and {Ru(phen)2}2+ moieties in acetonitrile. The charge-separated intermediate {Ru(III) (phen)2(pPDIp–•)} was detected by transient absorption spectroscopy upon electronic excitation in either the pPDIp or the complex moieties. The charge-separated intermediate species decays to generate the triplet state 3*pPDIp-Ru(II) (τP = 1.8 μs) that sensitizes the formation of singlet molecular oxygen with quantum yield ϕΔ = 0.57. The dyad in deaerated acetonitrile solutions is reduced by triethylamine (NEt3) to the [Ru(II) (phen)2(pPDIp•–)] radical anion in the dark. The electron-transfer reaction is accelerated by light absorption. By photolysis of the radical anion, a second electron transfer reaction occurs to generate the [Ru(II) (phen)2(pPDIp2–)] dianion. The changes of the color of solution indicate the redox states of compl...

Photodynamic properties and photoinactivation of Candida albicans mediated by brominated derivatives of triarylmethane and phenothiazinium dyes

The photodynamic activity of brominated derivatives of New Fuchsin and Azure B was studied in solution and in cell suspensions of Candida albicans. The spectroscopic and photodynamic properties of these photosensitizers were compared with those of Crystal Violet and Azure B, which represent active photosensitizer related to each family of compounds. Triarylmethane derivatives absorb intensely with a band centered at ∼ 570 nm, while the phenothiazinium dyes at ∼ 650 nm. Photooxidation of 9,10-dimethylanthracene was observed using phenothiazinium compounds indicating the formation of singlet molecular oxygen, while it was not detected using triarylmethane agents. However, triarylmethane dyes were able to photooxidize l-tryptophan. In yeast cell suspensions, the photosensitized inactivation of C. albicans increases with photosensitizer concentration, causing a ∼ 5 log decrease of cell survival, when the cultures are treated with 20 μM of Crystal Violet and irradiated for 60 min. Under these conditions, the photodynamic activity of 50 μM Azure B induced a ∼ 3 log decrease of cell survival. Studies of photodynamic action mechanism indicated that photoinactivation of C. albicans cells induced by triarylmethane compounds involves mainly type I photoprocess. Although, phenothiazinium derivatives produce singlet molecular oxygen, a contribution of other reactive oxygen species cannot be discarded in the photoinactivation of C. albicans.

Mechanism for the enhanced peroxidation of linoleic acid by a titanium dioxide/hypochlorite system

Nanosized titanium dioxide (TiO2) is a common component of sunscreen preparations and cosmetics as it reflects UV and visible light in accordance to Rayleigh’s law. However, in aqueous environments, TiO2 is an efficient photocatalyst, producing superoxide (O2-) and hydroxyl (HO) radicals, which are highly damaging to biomolecules. We investigated the role of TiO2 in promoting the peroxidation of linoleic acid (LA) alone and in the presence of hypochlorous acid (HOCl). TiO2 significantly enhanced peroxidation of LA, which was further enhanced in the presence of HOCl. This latter finding involved the formation of singlet molecular oxygen in a Russell-type mechanism appearing to involve preformed lipid hydroperoxides (LOOH). In addition to lipid peroxidation, HOCl also mediated formation of 18:1 monochlorohydrins, which in the presence of TiO2 appeared to decompose to kinetic products which supplemented peroxidation of linoleic acid. We present a theoretical mechanism which fits the available experimental data and may partially explain the dichotomy associated with HOCls role in lipid modification.

Photoprocesses of chlorin e6 glucose derivatives

The ground and excited state processes of chlorin e6, the monomethyl ester C1, the glucose derivative C2 and the 3-heptylchlorin-glucose C3, were studied in solvents of lower and higher polarity. The excited singlet and lowest triplet states of C1-C3 were characterized by spectroscopic methods for several conditions. The quantum yields of formation of singlet molecular oxygen and the other triplet properties of the three chlorins and chlorin e6 are similar, whereas the fluorescence quantum yield decreases on going from C1 to C3. Time-resolved optoacoustic experiments revealed a ca. 30 kJ mol(-1) higher triplet level for C3 with respect to C1/2.

Photogeneration of singlet molecular oxygen (1O2, 1Δ g ) using monomeric and aggregated forms of copper tetra-4-(morpholine-4-yl)-tetra-5-(2-naphthoxy)phthalocyanine in organic solvents

Luminescence and chemical acceptors methods were used to study the photosensitized formation of singlet molecular oxygen (1O2, 1Δ g ) by monomer and aggregated forms of copper tetra-4-(morpholine-4-yl)-tetra-5-(2-naphthoxy)phthalocyanine in benzene, benzene-d 6, acetone, DMF, and pyridine. The values of 1O2 quantum yield were determined. The effectiveness of the 1O2 photoproduction was shown to be reduced due mainly to the aggregation of the copper phthalocyanine molecules.

Photoreduction and Ketone‐sensitized Reduction of Alkaloids

The photoprocesses of berberine, palmatine, coralyne, sanguinarine, flavopereirine and ellipticine were studied in several solvents. The quantum yields Φ(Δ) of singlet molecular oxygen formation of berberine, palmatine and sanguinarine are moderate in dichloromethane (0.2-0.6) and much smaller in acetonitrile or trifluoroethanol. For the other alkaloids examined, Φ(Δ) is rather independent of solvent polarity. The direct and ketone-sensitized photolysis, using steady-state irradiation at 313 nm or 248/308 nm laser pulses, was studied by absorption and fluorescence spectroscopy. Thereby, radicals were observed yielding eventually dihydro derivatives as major products, which are thermally back-converted on admission of oxygen. The quantum yield of conversion of alkaloids to dihydroalkaloids is enhanced in the presence of triethylamine. The reaction in the presence of ketones and electron or H-atom donors has a quantum yield of close to unity.

Meso‐Indolo[3,2‐b]carbazolyl‐Substituted Porphyrinoids: Synthesis, Characterization and Effect of the Number of Indolocarbazole Moieties on the Photophysical Properties

Abstractmeso‐Indolocarbazolylporphyrins endowed with a different number of indolocarbazole units have been synthesized via condensation of an appropriately substituted monoformylated 5,11‐dihydroindolo[3,2‐b]carbazole precursor and mesityldipyrromethane. Under specific conditions, analogous meso‐indolocarbazolylcorroles could also be prepared. The photophysical features of the novel luminescent free‐base and Zn‐porphyrin derivatives were investigated. The introduction of indolocarbazole substituents results in progressive bathochromic shifts of the porphyrin absorbance and fluorescence bands due to the rising energy of the a2u orbital. The excitation energy is efficiently transferred from the meso‐indolocarbazole units to the porphyrin macrocycle. An increased number of indolocarbazole moieties does not lead to porphyrin fluorescence quenching; on the contrary, a small increase of the fluorescence quantum yield is observed. The main route for excitation energy deactivation of all the studied porphyrins is intersystem S1→T1 crossing, with the intersystem crossing quantum yield, as determined by the photosensitized formation of singlet molecular oxygen, being as high as about 70 % for the free‐bases and more than 80 % for the Zn complexes. The intersystem crossing quantum yield seems to be barely affected by meso‐indolocarbazole substitution. A noticeable part of the excitation energy was found to deactivate through radiationless internal S1→S0 conversion.

Formation of singlet molecular oxygen associated with the formation of superoxide radicals in aqueous suspensions of TiO2 photocatalysts

The production and decay of singlet molecular oxygen (1O2) in TiO2 photocatalysis were investigated by monitoring its phosphorescence under various reaction conditions. First, the effects of additives such as KBr, KSCN, KI, H2O2, and ethanol on the amount of 1O2 produced by photo excitation of P25 TiO2 were measured. The same additives were employed to investigate the effect on the amount of O2− produced. Comparison between the effects on 1O2 and O2− suggested that 1O2 is formed by the electron transfer mechanism, the reduction of molecular oxygens to O2− by photogenerated electrons and the subsequent oxidation of O2− to 1O2 by photogenerated holes. The formation of 1O2 decreased at pH < 5 and pH > 11, indicating that the intermediate O2− is stabilized at the terminal OH site of the TiO2 surface in the pH range of 5 < pH < 11. Eighteen commercially available TiO2 photocatalysts were compared on the formation of 1O2 and O2− in an aqueous suspension system. The formation of 1O2 was increased with decreasing size of TiO2 particles, indicating that a large specific surface area causes a higher possibility of reduction producing O2− and then a large amount of 1O2 is formed. The difference in the crystal phase (rutile and anatase) did not affect the formation of 1O2.

Photodimers of N-alkyl-3,4-dimethylmaleimides—Product ratios and reaction mechanism

The photodimerization of three N-alkyl-3,4-dimethylmaleimides (DMI) – frequently used to form crosslinks in polymers – was examined. N-Isobutyl-3,4-dimethylmaleimide, N-(2-acetamido)ethyl-3,4-dimethylmaleimide, N-(3-propionic acid)-3,4-dimethylmaleimide were studied by steady-state and time-resolved photolysis in various solvents. Unexpectedly, under all conditions a single-bonded dimer distinct from the well-known cyclobutane fused cyclic dimer was identified. Depending on the solvent the single-bonded dimer amounts to 5–50% of the product yield, allowing for considerable selectivity. Dimerization quantum yields, Φ dim, range from 0.01 to 0.5. Results from time-resolved experiments establish the involvement of the triplet state in the photodimerization mechanism, the quantum yield of singlet molecular oxygen formation accounts for the Φ dim values. Electron transfer and hydrogen abstraction reactions of DMI in polar and protic solvents, in contrast to N-alkylmaleimides, are inefficient processes.

Synthesis and photophysics of porphyrin–fullerene donor–acceptor dyads with conformationally flexible linkers

The synthesis and photophysics of a series of porphyrin–fullerene (P–C 60) dyads in which the two chromophores are linked by conformationally flexible polyether chains is reported. Molecular modeling indicates the two moieties adopt a stacked conformation in which the two chromophores are in close proximity. Photoexcitation of the free base dyads in polar solvents such as tetrahydrofuran and benzonitrile, causes electron transfer (ET) to generate charge-separated radical pair (CSRP) states, which were directly detected using transient absorption (TA) techniques. In nonpolar solvents such as toluene, where CSRP states were not directly detected, fullerene triplet state states were formed, according to TA studies as well as singlet oxygen sensitization measurements. The low value of the quantum efficiency for sensitized formation of singlet molecular oxygen [O 2( 1Δ g)] in toluene and chloroform indicates that singlet energy transduction to give H 2P– 1C 60*, followed by intersystem crossing to H 2P– 3C 60* and energy transfer to 3O 2, is not the operative mechanism. Rather, a mechanism is proposed involving ET to give CSRP states followed by exergonic charge recombination to eventually generate fullerene triplets. Such a mechanism has been demonstrated experimentally for structurally related P–C 60 dyads. For the corresponding ZnP–C 60 dyads with flexible linkers, only photoinduced ET to generate long-lived CSRP states is observed. Photoinduced charge separation in these dyad systems is extremely rapid, consistent with a through space rather than through-bond mechanism. Charge recombination is up to three orders of magnitude slower, indicating this process occurs in the inverted region of the Marcus curve that relates ET rates to the thermodynamic driving force. These observations once again demonstrate the advantages of incorporating fullerenes as electron acceptor components in photosynthetic model systems.

Direct evidence of singlet molecular oxygen [O2(1Deltag)] production in the reaction of linoleic acid hydroperoxide with peroxynitrite.

Peroxynitrite (ONOO-), a biologically active species, can induce lipid peroxidation in biological membranes, thereby leading to the formation of various hydroperoxides. We report herein on the formation of singlet molecular oxygen [O(2) ((1)Delta(g))] in the reaction of peroxynitrite with linoleic acid hydroperoxide (LAOOH) or (18)O-labeled LAOOH. The formation of O(2) ((1)Delta(g)) was characterized by (i) dimol light emission in the red spectral region (lambda > 570 nm) using a red-sensitive photomultiplier; (ii) monomol light emission in the near-infrared region (lambda = 1270 nm) with a liquid nitrogen-cooled germanium diode or a photomultiplier coupled to a monochromator; (iii) the enhacing effect of deuterium oxide on chemiluminescence intensity, as well as the quenching effect of sodium azide; and (iv) chemical trapping of O(2) ((1)Delta(g)) or (18)O-labeled O(2) ((1)Delta(g)) with the 9,10-diphenylanthracene (DPA) and detection of the corresponding DPAO(2) or (18)O-labeled DPA endoperoxide by HPLC coupled to tandem mass spectrometry. Moreover, the presence of O(2) ((1)Delta(g)) was unequivocally demonstrated by a direct spectral characterization of the near-infrared light emission attributed to the transition of O(2) ((1)Delta(g)) to the triplet ground state. For the sake of comparison, O(2) ((1)Delta(g)) deriving from the thermolysis of the endoperoxide of 1,4-dimethylnaphthalene or from the H(2)O(2)/hypochlorite and H(2)O(2)/molybdate systems were also monitored. These novel observations identified the generation of O(2) ((1)Delta(g)) in the reaction of LAOOH with peroxynitrite, suggesting a potential O(2) ((1)Delta(g))-dependent mechanism that contributes to cytotoxicity mediated by lipid hydroperoxides and peroxynitrite reactions in biological systems.

Direct evidence of singlet molecular oxygen [O 2[formula omitted]] production in the reaction of acetonitrile with hydrogen peroxide in alkaline solutions

In this work, we report direct evidence of the formation of singlet molecular oxygen [O 2 ( 1 Δ g ) ] in the reaction of hydrogen peroxide (H 2O 2) with acetonitrile in alkaline solutions. The formation of O 2 ( 1 Δ g ) was characterized by: (i) the dimol light emission in the red spectral region (>610 nm) using a red-sensitive photomultiplier tube (PMT); (ii) the monomol light emission in the near-infrared region (1270 nm) with a newly available tube coupled to a monochromator; and (iii) the quenching effect of sodium azide. Direct spectral characterization of the near-infrared emission that attributes the emission to the transition of O 2 ( 1 Δ g ) to the triplet ground state (O 2 ( 3 Σ g −) ) was done to unequivocally demonstrate the presence of O 2 ( 1 Δ g ) . For comparison, O 2 ( 1 Δ g ) derived from the thermolysis of the endoperoxide of 1,4-dimethylnaphthalene or from the H 2O 2/hypochlorite system were also monitored. The product of the reaction (acetamide) was also detected by mass spectrometry (MS). This evidence clearly demonstrates that the reaction of H 2O 2 with acetonitrile in alkaline solutions generates O 2 ( 1 Δ g ) , confirming the mechanism proposed by McKeown and Waters.

Formation of singlet molecular oxygen by the Radziszewski reaction between acetonitrile and hydrogen peroxide in the absence and presence of ketones

Measurements of the infrared phosphorescence of singlet molecular oxygen (1O2) at 1270 nm have been used to demonstrate the formation of 1O2 by the Radziszewski reaction between acetonitrile and hydrogen peroxide. The kinetics of the Radziszewski reaction either alone or in the presence of ketones have been studied by this technique. The rate-determining step of the 1O2 formation of the reaction in the absence of ketones was found to be independent of both the concentration of acetonitrile and that of hydrogen peroxide. The kinetic data, the results of the volumetric measurements of the oxygen liberated and the results of the determination of the amount of 1O2 generated by the reaction are consistent with the assumption that the reaction between acetonitrile and hydrogen peroxide occurs via the heterolytic decomposition of the intermediate, peroxyacetimidic acid (PAIA), forming 1 mol acetamide and 0.5 mol 1O2 according to the stoichiometric equation: CH3CN + H2O2 → CH3C(O)NH2 + 0.5 1O2. The rate constant of the heterolytic decomposition of PAIA was determined to be k8 = 1.2 × 10−3 dm3 mol−1 s−1 at T = 30 °C. From the measurements at different pH values in the range 9.1 < pH < 12.0 the pKa(PAIA) value was estimated to be 11.1 at T = 30 °C. The investigation of the reaction between acetonitrile and hydrogen peroxide by using N,N-dimethyl-4-oxopiperidinium nitrate as catalyst, has unequivocally shown that the rate of 1O2 formation is considerably enhanced by this ketone. For the ketone-catalysed decomposition of PAIA a rate law can be derived showing a first order dependence on the concentration of acetonitrile and hydrogen peroxide at a given pH. In accordance with the observed rate law are the results with acetonitrile in 50% acetone containing a tenfold excess of hydrogen peroxide at pH 8.2 and T = 60 °C.

Photophysical and Photochemical Processes of 2-Methyl, 2-Ethyl, and 2-tert-Butylanthracenes on Silica Gel. A Substituent Effect Study

The photophysics and photochemistry of 2-methylanthracene (2MA), 2-ethylanthracene (2EA), and 2-tert-butylanthracene (2TBA) adsorbed on silica was studied at a silica/air interface to determine the substituent effect on the above processes. In contrast to anthracene (AN), which forms ground state stable pairs at surface coverages as low as 1% of a monolayer, 2MA, 2EA, and 2TBA show no evidence of any ground state pairing even at high surface coverages. Diffuse reflectance and fluorescence data indicate that 2MA, 2EA, and 2TBA crystallize on the surface at higher surface coverages. Photolysis of 2MA, 2EA, and 2TBA at a silica/air interface proceeds more efficiently than photolysis of AN to produce the corresponding 9,10-endoperoxides formed by the addition of singlet molecular oxygen (type II) to the ground state molecules. Thermally unstable endoperoxides slowly decompose on silica surface to give the corresponding 9,10-quinones and other hydroxylated products of 2MA, 2EA, and 2TBA. Although photolyzed AN/silica samples show a significant amount of dimer formation at low surface coverages, no evidence of any dimer is observed in photolyzed samples of 2MA, 2EA, and 2TBA at low surface coverages. Small amount of dimers (isomeric), however, are observed in the photolyzed samples of 2MA, 2EA, and 2TBA at higher surface coverages suggesting that the crystal forms of these molecules may be involved in the dimerization process. The photolysis rate decreases as the surface coverage is increased. This behavior can be attributed to the involvement of crystals (formed at higher surface coverage) which act as a light sink to absorb the incident light without producing any net chemistry. The photolysis rate decreases by an order of magnitude in the presence of ≤1 monolayer of physisorbed water. Time-resolved transient studies of excited triplet states of 2MA, 2EA, and 2TBA have revealed that triplet lifetimes are shortened on wet silica. Furthermore, the efficiency of singlet molecular oxygen formation drops significantly on wet silica. These results suggest that the decrease in photolysis rate is due to lowering of the singlet oxygen quantum yield in the presence of physisorbed water.

Coupling photochemical reaction detection based on singlet oxygen sensitization to capillary electrochromatography

Despite the impressive separation efficiency afforded by capillary electrochromatography (CEC), the detection of UV-absorbing compounds following separation in capillary dimensions remains limited by the short path length (5-75 microm) through the column. Moreover, analytes that are poor chromophores present an additional challenge with respect to sensitive detection in CEC. This paper illustrates a new photochemical reaction detection scheme for CEC that takes advantage of the catalytic nature of type II photooxidation reactions. The sensitive detection scheme is selective toward molecules capable of photosensitizing the formation of singlet molecular oxygen (1O2). Following separation by CEC, UV-absorbing analytes promote groundstate 3O2 to an excited state (1O2) which reacts rapidly with tert-butyl-3,4,5-trimethylpyrrolecarboxylate, which is added to the running buffer. Detection is based on the loss of pyrrole. The reaction is catalytic in nature since one analyte molecule may absorb light many times, producing large amounts of 1O2. The detection limit for 9-acetylanthracene, following separation by CEC, is approximately 6 x 10(-9) M (S/N = 3). Optimization of the factors effecting the S/N for four model compounds is discussed.

Participation of the photosensitizer alpha-terthienyl in the peroxidase-catalyzed oxidation of indole-3-acetic acid.

The plant photosensitizer alpha-terthienyl (alpha T) is toxic toward a variety of organisms, and normally requires exposure to ultraviolet-A radiation for activation and singlet molecular oxygen formation. However, some toxicity has also been reported to occur in the dark. One hypothesis that has been proposed to account for this light-independent toxicity is that the sensitizer becomes activated by energy transfer from the excited-state products of enzymatic reactions. We have investigated this hypothesis using the horseradish peroxidase (HRP)-catalyzed oxidation of indole-3-acetic acid (IAA), which generates indole-3-aldehyde in an excited triplet state. Light is emitted during the IAA/HRP reaction at acidic pH, is increased by inclusion of alpha T and is not observed with heat-denatured HRP. The rates of both the oxidation of IAA and the subsidence of light emission are more rapid in the IAA/alpha T/HRP system than with IAA and HRP alone, indicating that the presence of alpha T accelerates the reaction. Bleaching occurs at the wavelength of maximal alpha T absorbance and is promoted by the inclusion of IAA. Readdition of both IAA and alpha T to a spent reaction mixture is required to restore light emission after it has subsided, further suggesting that both are consumed in the reaction. We were unable to detect measurable quantities of singlet molecular oxygen formation in this system. These results do not support the energy transfer hypothesis, but instead are more compatible with a model proposed by Krylov and Chebotareva [Krylov, S. N. and A. B. Chebotareva (1993) FEBS Lett. 324, 6-8] for the co-oxidation of IAA and xanthene dyes.