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Exosomes are important intercellular communication vehicles, secreted into body fluids by multiple cell types, including tumor cells. Exosomes stimulate angiogenesis, tumor cell migration, and invasion, as was proved in cell culture studies. Recently, it was found that some of the exosomes circulating in the blood are associated with the surface of blood cells (exosome-assCS), but their role in the dissemination of the tumor process remains unclear. We performed mass spectrometry analysis of proteomes from isolated plasma exosomes and blood cells associated exosomes of healthy females (HFs) and breast cancer patients (BCPs). Exosomes were separated using ultrafiltration and ultracentrifugation approaches; and their origin was verified using transmission electron microscopy and flow cytometry. Gene Ontology (GO) Functional Enrichment analysis via FunRich software showed that proteins previously detected in the cytoplasm, Oxford comma nucleus are predominant in the composition of plasma exosomes and exosomes-assCS. In breast cancer, a redistribution of the functions of plasma exosomes and blood cell-associated exosomes has been established: the proportion of transport, catalytic, and motor protein increases in plasma exosomes, and, conversely, the proportion of proteins that regulate transcriptional activity decreases in blood cell-associated exosomes. Also, the proportion of proteins involved in signal transduction and cellular communication decreases and the proportion of proteins involved in protein metabolism increases in both exosome fractions in the blood of breast cancer patients. Using the dbDEPC 3.0 database (database of Differently Expressed Proteins in Human Cancer), it was shown that 64% of the proteins of blood cell associated exosomes from BCPs are of tumor origin. Profiling of exosomal blood proteins using QuickGO annotations showed that the proportions of BCPs blood exosomal proteins involved in epithelial-mesenchymal transition (EMT), cell motility, invasion, and immune response are comparable in plasma exosomes and blood cell-associated exosomes. However, the proportion of proteins inhibiting these processes is 1.6–4 times lower in BCPs exosomes than in the blood exosomes of HFs. Thus, conducted proteome analysis of the blood cell associated exosomes in patients with breast cancer highlights the role of these exosomes in the dissemination of the tumor process along with plasma exosomes.

Dependences of the fluorescence and triplet state quantum yields of kynurenic acid (1) and kynurenine yellow (2) in water—glycerol mixtures on medium viscosity have been studied. The main channel of the singlet excited state decay of compound 1 is the intersystem crossing, which rate weakly depends on the viscosity; only a small (approximately 1.5-fold) increase in the fluorescence yield was found for this compound with the increase of the solution viscosity from 0.84 cP (aqueous solution) to 78 cP (86% glycerol). The deactivation of the S1 state of compound 2 is caused mainly by the internal conversion, and a noticeable increase of the fluorescence yield (approximately 3-fold), as well as the change in the photolysis product yields, was observed with the increasing percentage of glycerol in the mixture. The triplet state quantum yields for compounds 1 and 2 remained unchanged with the variation of the glycerol content in the mixture.

Microbial alkaloid prodigiosin is proposed as a new effective matrix for the analysis of low-molecular- weight plant antioxidants by MALDI mass spectrometry in the negative ion detection mode. It is demonstrated that the physicochemical characteristics of prodigiosin, such as ability to activate ionization/ desorption of analyte, high absorption coefficient at the working wavelengths of lasers, good solubility in solvents used for analytes, uniformity of crystallization on the target, stability under working conditions, and the absence of interfering peaks of fragment and cluster ions, are consistent with the recommendations for effective matrices. Prodigiosin as a matrix ensures the high-resolution detection of various plant antioxidants in complex mixtures. The obtained mass spectra of flavonoid aglycones were characterized by high quality (signal-to-noise ratio higher than 103) and reproducibility. The use of prodigiosin for analyzing a complex preparation of plant antioxidants ensures the simultaneous recording and interpretation of more than two dozens of ions corresponding to various antioxidants with the molecular weights of 100–500 Da.

The reactions of photoexcited kynurenic acid (KNA) with bovine α-crystallins under anaerobic conditions proceed via the electron transfer from tryptophan (Trp) and tyrosine (Tyr) residues to the triplet KNA molecules. The subsequent radical reactions lead to the protein aggregation and insolubilization. The absorption of the photolyzed proteins at 335 nm as well as their total fluorescence significantly increases, while the tryptophan-related fluorescence decreases. It has been established that the alterations of the protein optical properties are related to the modifications of Trp residues. Intrinsic lens antioxidants ascorbate (Asc) and glutathione (GSH) that are present in the human lens at the millimolar level effectively block the formation of the observed light-induced protein modifications. The protective effect of Asc was attributed to its ability to quench highly reactive triplet states, while the role of GSH, most likely, corresponds to the reduction of photochemically formed radicals into a diamagnetic state. The results obtained disclose the possible mechanism of UVA-induced modifications of the lens crystallins, leading to the formation of cataract, and the role of major lens antioxidants Asc and GSH in the protection of the lens proteins.

The ability of SkQ1 eye drops to slow down the cataract development is demonstrated on the senescence-accelerated OXYS rats: the SkQ1 treatment leads to the considerable improvement of the lens condition as compared to the control group. The comparison of the chaperone activities of α-crystallins isolated from the rat lenses did not reveal significant difference between SkQ1-treated and control rats. The contents of major metabolites (23 compounds) in lenses of SkQ1-treated and untreated rats are also very similar, though the concentration of reduced glutathione (GSH) in lenses of SkQ1-treated rats is 12% lower. This difference may be attributed to the reduction of the oxidative stress under action of SkQ1 eye drops, and to the decreased requirement to produce high amounts of this antioxidant.

Dynamics of transformations of excited states and active transient species generated in the photolysis of nitrated 1,2-dihydroquinolines (N-DHQ) has been studied by femto- and nanosecond laser pulse photolysis. Spectral and kinetic parameters of primary photophysical and photochemical processes have been determined, and their dependence on the substituent position at the aromatic ring of 1,2-dihydroquinoline (DHQ) and on the wavelength of excitation light has been established. The lifetime of the excited singlet state S1 in N-DHQ is ca. 100 and 500 fs for 8- and 6-nitro-substituted DHQ, respectively, which is shorter in comparison with DHQ without the nitro group by a factor of 104 and more. The major decay channel of the S1 state is the successive formation of three transient species with lifetimes of 0.5 to 16 ps. A triplet state is generated only upon excitation of the short-wavelength band by UV light. The quantum yield of the triplet state depends on the structure of N-DHQ.

Supplementation of senescence-accelerated OXYS rats with the mitochondria-targeted antioxidant SkQ1 and with the powder from Cistanche deserticola results in the deceleration of the cataract development and even in the improvement of lens transparency. The therapeutic effect of these preparations correlates with a significant elevation of tryptophan and kynurenine levels in the lens. This finding is attributed to a deceleration of the tryptophan and kynurenine oxidation due to antioxidant-assisted reduction of oxidative stress in the lens.

The reactions of triplet kynurenine and 4-(2-aminophenyl)-4-oxocrotonic acid, formed upon the thermal decomposition of kynurenine, with nitroxyl radicals and cyclic N-hydroxylamines were studied. Nitroxyl radicals were found to quench efficiently the triplet state of kynurenine (rate constant 3–6·.108 L mol-1 s-1). The quenching proceeds via the spin-exchange mechanism and affords no new products. Neither nitroxyl radicals, nor hydroxyl-lamines react with 4-(2-aminophenyl)-4-oxocrotonic acid under conditions similar to physiological.

Quantum yields of photodecomposition and triplet state formation under aerobic and anaerobic conditions are determined for kynurenine (KN), 3-hydroxykynurenine (3OHKN), xanthurenic acid (XAN), and kynurenine adducts of glutathione (GSH-KN), cysteine (Cys-KN), histidine (His-KN), and lysine (Lys-KN) in aqueous solutions. The highest yields of anaerobic photodecomposition were obtained for GSH-KN and His-KN adducts, which correlates with the highest triplet yields for these compounds. In aerobic conditions, the photodecomposition yields for all compounds under study increase; the highest decomposition rates were observed for His-KN and 3OHKN. The fast decomposition of the latter is attributed to the dark autoxidation of the starting compound.

Kynurenine and its derivatives are present in the lens of the human eye, where they perform the function of UV filters. Photochemical and thermal reactions of these compounds, which can result in irreversible modification of the lens proteins, and play an important role in the cataract development, are considered. Bibliography - 123 references.

Photochemical reactions involving kynurenines, viz., molecules present in the eye lens, can result in modifications of the lens proteins and cause a development of a cataract. The rate constants of the reactions of photoexcited kynurenine with several amino acids and antioxidants contained in the lens were measured. The most efficient quenchers of triplet kynurenine are amino acids tryptophan and tyrosine, as well as antioxidant ascorbate. In all cases, the quenching reaction proceeds by the electron transfer mechanism, except for the reaction with oxygen where transfer of the triplet energy to the oxygen molecule occurs.

The photoionization of kynurenine (KN) and 3-hydroxykynurenine (3OHKN) in aqueous solutions proceeds via a biphotonic mechanism. The precursors for ionization are the triplet states TKN and T3OHKN, absorbing the second light quantum. The addition of solvated electrons to the parent molecules with the rate constant 2.0 × 1010 M−1 s−1 results in the formation of anion radicals, the latter in neutral solutions undergoes fast protonation. The individual spectra of all intermediates – triplet states, cation radicals, electron adducts – formed under UV irradiation of KN and 3OHKN are obtained.

Condensation of formaldehyde and lower carbohydrates (glycolaldehyde, glyceraldehyde, and dihydroxyacetone) is effectively catalyzed by heterogeneous and homogeneous phosphates in neutral aqueous medium. The interaction of formaldehyde and dihydroxyacetone leads to the preferential formation of 3-pentulose and erythrulose with yields of 40% and 45%, respectively. In absence of formaldehyde, the condensation of glycolaldehyde and glyceraldehyde catalyzed by phosphates leads to the formation of ribose and fructose. We demonstrate the possibility of formation of higher monosaccharides from pure formaldehyde in the course of the combined photochemical and phosphate-catalyzed reactions in plausible prebiotic conditions.

Pyrene fluorescence quenching by phenylazide derivatives with donor and acceptor substituents has been studied by fluorescence spectroscopy and flash photolysis. The rate constants of quenching (k q) in acetonitrile ((0.2–1.2) × 1010 l mol−1 s−1) are found to be close to a diffusion limit; the rate constants were somewhat higher for perfluoro-substituted arylazides. It is found that k q does not depend on solvent polarity; the formation of the pyrene cation in the course of pyrene fluorescence quenching by tolylazide was not detected. Pyrene fluorescence quenching occurred by an energy-transfer mechanism; this is supported by the coincidence of the quantum yields of the direct and sensitized photodecomposition of tolylazide. As estimated, energy transfer in rigid media occurs at characteristic distances of about 10 A.

Triplet tryptophanTTrpH quenching the by nucleotides adenosine-5′-monophosphate (AMP), cytidine-5′-monophosphate (CMP), guanosine-5′-monophosphate (GMP), and uridine-5′-monophosphate (UMP) is reported for the first time. The quenching rate constants for AMP, CMP, and UMP are of the same orderkq=(3–6)·108 M−1s−1, for GMP this value is 5.6·107 M−1s−1. This difference correlates with the reduction potentials of the nucleotides under study, pointing at the involvement of the electron transfer process.TTrpH quenching by AMP, CMP, and GMP does not result in the radical formation, whereas in the reaction ofTTrpH with UMP radicals are formed with the quantum yield Φ≈0.13, the, radical formation is confirmed by both laser flash photolysis and time-resolved chemically induced dynamic nuclear polarization measurements.

Intramolecular electron transfer (IET) from tyrosine to tryptophan cation radicals is investigated using time resolved chemically induced dynamic nuclear polarization (CIDNP) spectroscopy in combination with laser flash photolysis. In both the tryptophan-tyrosine dipeptide and the denatured state of hen lysozyme in aqueous solution, the transformation TrpH+. → TyrO. by IET leads to an increase in the tyrosine radical concentration, growth in the tyrosine CIDNP signal, fast decay of the tryptophan CIDNP, and inversion of the phase of the CIDNP of the photosensitizing dye, 2,2′-dipyridyl. IET effects are not observed for mixtures of the amino acid or for the native state of lysozyme. The steady state CIDNP effects seen for denatured lysozyme thus depend not only on the accessibility of the amino acid residues on the surface of the protein but also on the reactivity of the radical intermediates.

The rate constant k CO of decarbonylation of phenylacetyl radicals generated by photolysis of dibenzyl ketone was measured by laser flash photolysis technique in six solvents in a wide temperature range. The pre-exponential factors A and activation energies E a of decarbonylation were found for all solvents. The k CO value decreases with an increase in the dielectric constant e of the solvent, whereas an increase in the ability of the solvent for hydrogen bonding increases k CO. The results of quantum-chemical calculations confirm the mutual compensation of the contributions of specific and nonspecific solvations to the activation energy of decarbonylation in alcohols.

The kinetics and mechanisms of the reactions of singlet perfluoro-4-biphenylnitrene and N -propyl- 4-nitreno-2,3,5,6-tetrafluorobenzylamide with various amines, pyridine, and dimethylsulfoxide were studied by laser flash photolysis. The reactions of singlet arylnitrenes with amines are two-step processes. The primary step of the process is adduct formation; the rate constant of this reaction is high and lies within the range 4 〈 10 7 -2 〈 10 8 l mol -1 s -1 for the tested secondary amines. The second step (1,2-hydrogen shift) was acceler- ated in the presence of water.

The kinetics of the nuclear polarization formed during the photolysis of acetone in isopropanol- d 8 were analyzed quantitatively. Model calculations show that the spin-selective recombination of radicals gives rise to the electron polarization and, with regard to the electron-nuclear cross-relaxation, are adequate to describe the formation of the net nuclear polarization of the reaction products even if the solution contains only one type of radical. For the 2-hydroxy-2-propyl radicals at a magnetic field of 7 T, fitting the theoretical results to the experimental data gives the electron relaxation time T 1 e = 1.0 ± 0.2 μ s and the cross-relaxation time T x = 92 ± 18 μ s.

[EN] The intramolecular electron transfer in the naproxen-N-methylpyrrolidine dyad has been investigated by spin chemistry methods. The existence of CIDNP in a high magnetic field points to electron transfer as a possible mechanism of the quenching of the excited state of a dyad. However, the failure to detect magnetic field effects on triplet yield makes us conclude that this quenching mechanism is not the only one. The observation of CIDNP effects in the dyad in the media of low polarity and the short risetime of triplet state formation indicate a potential role of exciplex in the quenching of the excited state of the dyad., This work was supported by the Grants 08-03-00372 and 11-03-01104 of Russian Foundation of Basic Research, and the grant of Priority Programs of RAS, No. 5.1.5.

The photolysis of aliphatic ketones using laser pulse excitation (λ = 308 nm) was investigated by the time-resolved flash chemically induced dynamic nuclear polarization (CIDNP) method. The contributions of geminate and homogeneous processes to the CIDNP effects were separated. It was shown that the lifetimes of the triplet molecules can be determined from an analysis of CIDNP formation kinetics and from the Stern—Volmer dependence of the amplitude of geminate polarization in the presence of quencher (piperylene). Irradiation of the samples with high-intensity laser light revealed the presence of two-photon processes which are demonstrated by a change in the correlation of CIDNP line intensities of different products with variations in laser radiation energy and by the non-linear character of the Stern—Volmer dependence. Increased irradiation intensity leads to the appearance of radicals resulting from the decomposition of highly excited triplet molecules of aliphatic ketones. Model calculations of CIDNP kinetics and the Stern—Volmer dependence were performed in terms of a kinetic scheme involving short-lived triplet states of aliphatic ketones. The calculations enable an estimation to be made of the contribution of two-photon processes to the photolysis and the molar extinction coefficients of the triplet state for methylethylketone and cyclododecanone.

Submicrosecond flash-CIDNP was used to study the dependences of the kinetics and amplitude of CIDNP, arising in biradicals produced in the photolysis of cyclic aliphatic ketones on the temperature and viscosity of the medium. It has been found that for biradicals the geminate CIDNP increases with the diffusion coefficient of the medium. The theoretical consideration of geminate recombinaiion kinetics is based on the numerical solution of the stochastic Liouville equation for the Fourier transform of the spin density matrix. The dependences of nuclear polarization on the lifetime of the triplet state of initial ketone, on electronic paramagnetic relaxation times, on scavenging rate, and on exchange interaction have been analyzed and compared with erperimental data. The conditions of quantum beats manifestaiion in CIDNP kinetics have been considered theoretically.

Kynurenine and its derivatives are present in the lens of the human eye, where they perform the function of UV filters. Photochemical and thermal reactions of these compounds, which can result in irreversible modification of the lens proteins, and play an important role in the cataract development, are considered. Bibliography - 123 references.

CIDNP with microsecond time resolution was used to study the kinetics of nuclear polarization formation during the photolysis of acetone in isopropanol over a wide temperature range. The coincidence of signs of multiplet effects of CIDNP (A/E) for cage and escape reaction products as well as the presence of net polarization (E) changing its sign with decreasing temperature have been revealed. The account of scalar electron-nuclear cross-relaxation has been shown to be necessary for the interpretation of the results obtained.

The influence of polarity and viscosity of medium on the geminate radical pairs formed by photoreaction of anthraquinone with triethylamine has been studied by time-resolved CIDNP and stimulated nuclear polarization (SNP). Radical and radical ion mechanisms have been shown to be the main ways of the reaction proceeding in non-polar and polar media, respectively. In media with intermediate polarity both the mechanisms are realized.

Laser flash photolysis of perfluoro-4-biphenyl azide andN-propyl-4-azido-2,3,5,6-tetrafluorobenzylamide in acetonitrile, water-acetonitrile mixtures, and HCl-containing solutions was studied. The absorption spectra of primary intermediates, singlet arylnitrenes(2a(S) and2b(S), respectively), were recorded. The absolute rate constants of their intersystem crossing in MeCN were measured, and the corresponding Arrhenius parameters were found from the temperature dependences of the rate constants of isomerization of singlet arylnitrenes2a(S) and2b(S) to azirines. Protonation of singlet arylnitrenes2a(S) and2b(S) was observed, the rate constants of their protonation were measured, and the transient absorption spectra of arylnitrenium ions were recorded. It was shown by quantum-chemical calculations (the hybrid B3LYP method) that the arylnitrenium ions that formed have the singlet ground state and the singlet-triplet gap is ∼20 kcal mol−1.

The quantum yield of photoionization of TrpH and IndH from the nonrelaxed prefluorescent state S* increases with the temperature decrease. This effect is attributed to the competition between temperature independent ionization and ultrafast thermal relaxation S* --> S1. The rate constant of the relaxation does not depend on the solvent and on the presence of the amino acid side chain: the temperature dependences of photoionization quantum yield, obtained for TrpH and IndH in different solvents, practically coincide. The activation energy for the relaxation rate constant Er approximately 4.5 kJ/mol probably corresponds to intramolecular process or to the formation of the vibronically excited transient complex between photoexcited molecule and solvent molecules.

Chemical reactions between the photoexcited triplet state of flavin mononucleotide and the aromatic amino acids, N-acetyl tryptophan (TrpH), N-acetyl tyrosine (TyrOH), and N-acetyl histidine (HisH) in aqueous solution have been studied in the pH range 2-12. Across the whole pH range, the principal mechanism of reaction of both TrpH and TyrOH is shown to be electron transfer. For HisH, the mechanism and rate of the reaction depend on the protonation state of the reactants. In acidic conditions (pH < 4), reaction does not occur. At 4 < pH < 11, the reaction proceeds via hydrogen atom abstraction with a rate constant varying from 3.0 x 10(6) to 2.5 x 10(8) M(-1) s(-1). In extremely basic solution (pH > 12) the mechanism switches to electron transfer.

The influence of the surfactants sodium dodecyl sulphate, cetyltrimethyl-ammonium bromide and triton X-100 on the photochemically induced dynamic nuclear polarization (CIDNP) of N-acetyl tyrosine has been investigated. Three photosensitizers were used to generate polarization: thionin, eosin Y and flavin mononucleotide. 600 MHz 1H photo-CIDNP experiments, supported by laser flash photolysis transient absorption measurements, indicate that the neutral triton surfactant has no influence on the nuclear polarization, but that the other two, charged, amphiphiles affect the photochemistry in a variety of ways, depending on the surfactant concentration and the identity of the sensitizer.

The singlet-triplet evolution of alkyl-acyl biradicals under S-T 0 mixing within submicrosecond time range has been studied by the method of flash CIDNP 1 H during the photolysis of cyclic aliphatic ketones. The maximum of nuclear polarization has been detected 160 ns after exciting laser pulse. The invariance of the CIDNP time-dependence curve relative to the isotopic substitution of ketone in the α-position at the polymethylene chain length of 10 and 11 has been demonstrated. The nuclear polarization evolution has been calculated both in terms of formal kinetics and using the kinematic approximation in the theory of geminate recombination for the two-position biradical model.

We have studied the dependence of CIDNP effects on the concentration of the starting material in photolysis of a solution of benzaldehyde in cyclohexane-D12 and in benzene-D6. Using an excimer laser (λ=308 nm), we have investigated the time evolution of the CIDNP signals with time resolution 1 μsec. We have determined the rate constant k1′ for transfer of a hydrogen atom from the hydroxybenzyl radical to the ground-state benzaldehyde molecule and also the rate constant for recombination of two hydroxybenzene radicals.

The anomalously large nuclear polarization during photolysis of cyclic ketones due to chemically induced dynamic nuclear polarization was observed. Nuclear polarization in such reactions may be used for creating a polarized target.(AIP)

Photolysis of acetone in the presence of various hydrogen donors and of 3-hydroxy-3-methyl-2-butanone involves the formation of propan-2-olyl radicals which show both electron and nuclear spin polarization. The electron polarization of the radicals leads to additional nuclear polarization of the reaction products. Transfer of electron to nuclear polarization can occur by cross-relaxation and cross-correlation. The latter is described in detail. Experimentally, the mechanisms lead to formation of net nuclear polarization for symmetrical radical pairs as well as an unusual kinetic behavior of multiplet effects