Quinone sensitized electron transfer photooxidation of nucleic acids: chemistry of thymine and thymidine radical cations in aqueous solution

The 2-methyl-1,4-naphthoquinone (MQ) sensitized photooxidation of nucleic acid derivatives has been studied by laser flash photolysis and steady state methods. Thymine and thymidine, as well as other DNA model compounds, quench triplet MQ by electron transfer to give MQ radical anions and pyrimidine or purine radical cations. Although the pyrimidine radical cations cannot be directly observed by flash photolysis, the addition of N,N,N',N'-tetramethyl-1,4-phenylenediamine (TMPD) results in the formation of the TMPD radical cation via scavenging of the pyrimidine radical cation. The photooxidation products for thymine and thymidine are shown to result from subsequent chemical reactions of the radical cations in oxygenated aqueous solution. The quantum yield for substrate loss at limiting substrate concentrations is 0.38 for thymine and 0.66 for thymidine. The chemistry of the radical cations involves hydration by water leading to C(6)-OH adduct radicals of the pyrimidine and deprotonation from the N(1) position in thymine and the C(5) methyl group for thymidine. Superoxide ions produced via quenching of the quinone radical anion with oxygen appear to be involved in the formation of thymine and thymidine hydroperoxides and in the reaction with N(1)-thyminyl radicals to regenerate thymine. The effects of pH were examined in the range pH 5-8 in both the presence and absence of superoxide dismutase. Initial C(6)-OH thymine adducts are suggested to dehydrate to give N(1)-thyminyl radicals.     

Reactivity of substituted charged phenyl radicals toward components of nucleic acids

Reactions of differently substituted phenyl radicals with components of nucleic acids have been investigated in the gas phase. A positively charged group located meta with respect to the radical site was employed to allow manipulation of the radicals in a Fourier-transform ion cyclotron resonance mass spectrometer. All of these electrophilic radicals react with sugars via exclusive hydrogen atom abstraction, with adenine and uracil almost exclusively via addition (likely at the C8 and C5 carbons, respectively), and with the nucleoside thymidine by hydrogen atom abstraction and addition at C5 in the base moiety (followed by elimination of (*)CH(3)). These findings parallel the reactivity of the phenyl radical with components of nucleic acids in solution, except that the selectivity for addition is different. Like HO(*), the electrophilic charged phenyl radicals appear to favor addition to the C5-end of the C5-C6 double bond of thymine and thymidine, whereas the phenyl radical preferentially adds to C6. The charged phenyl radicals do not predominantly add to thymine, as the neutral phenyl radical and HO(*), but mainly react by hydrogen atom abstraction from the methyl group (some addition to C5 in the base followed by loss of (*)CH(3) also occurs). Adenine appears to be the preferred target among the nucleobases, while uracil is the least favored. A systematic increase in the electrophilicity of the radicals by modification of the radicals' structures was found to facilitate all reactions, but the addition even more than hydrogen atom abstraction. Therefore, the least reactive radicals are most selective toward hydrogen atom abstraction, while the most reactive radicals also efficiently add to the base. Traditional enthalpy arguments do not rationalize the rate variations. Instead, the rates reflect the radicals' electron affinities used as a measure for their ability to polarize the transition state of each reaction.     

Coordination‐Induced Spin‐State Switching of an Aminyl‐Radical‐Bridged Nickel(II) Porphyrin Dimer between Doublet and Sextet States

A bis(Ni^II‐porphyrinyl)aminyl radical with meso‐C₆F₅ groups was prepared as a spin‐delocalized stable aminyl radical with a doublet spin state. Upon addition of pyridine, both Ni^II centers became hexacoordinated by accepting two axial pyridines, which triggered a spin‐state change of the Ni^II centers from diamagnetic (S=0) to paramagnetic (S=1). The resulting high‐spin Ni^II centers interact with the aminyl radical ferromagnetically to give rise to an overall sextet state (S=5/2). Importantly, this coordination‐induced spin‐state switching can be conducted in a reversible manner, in that washing of the high‐spin radical with aqueous hydrochloric acid regenerates the original doublet radical in good yield.     

Photochemistry of (η5)-C5H5)Mn(CO)2[C(C6H5)2]. Generation of the diphenylcarbene radical anion

The electronic absorption spectrum of (η⁵-C₅H₅)Mn(CO)₂[C(C₆H₅)₂]shows an intense maximum which is assigned to a MLCT transition in which the empty p_π orbital on the carbene carbon is populated. Upon irradiation of this band, the complex undergoes a decomposition with a disappearance quantum yield Φ = 0.10 ± 0.01 independent of solvent. In the CT excited state, the complex can be roughly described as containing d⁵ Mn^II and a diphenylcarbene radical anion ligand C(C₆H₅)₂^?. Due to the kinetic lability, the complex decomposes producing a Mn^II species and the free carbene radical anion, which then undergoes secondary reactions. In addition, small amounts of substitution product are observed. It is proposed that prior to total decomposition of the excited state, a radical pair (η⁵-C₅H₅)Mn(CO)₂S⁺/C(C₆H₅)₂^?forms (S = solvent). A back electron transfer from C(C₆H₅)₂^?to the labile cation competes with decomposition to produce the substituted complex and free carbene.     

Radical addition reactions: factors determining the transition-state geometry

Interatomic distances in the reaction centers of the addition reactions of (i) H^· to the C=C, C=O, N≡C, and C≡C bonds, (ii) ^·CH₃ radical to the C=C, C=O, and C≡C bonds, and (iii) alkyl, aminyl, and alkoxyl radicals to olefin C=C bonds were determined using a new semiempirical method for calculating transition-state geometries of radical reactions. For all reactions of the type X^· + Y=Z → X— Y—Z^· the r ^# _X...Y distance in the transition state is a linear function of the enthalpy of reaction. Parameters of this dependence were determined for seventeen classes of radical addition reactions. The bond elongation, Δr ^# _X...Y, in the transition state decreases as the triplet repulsion, electronegativity difference between the atoms X and Y in the reaction center, and the force constant of the attacked multiple bond increase. __________ Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 4, pp. 894–902, April, 2005.     

Photochemical generation and reactivity of the major hydroxyl radical adduct of thymidine

5,6-Dihydro-5-hydroxythymidin-6-yl radical (1), the major reactive intermediate resulting from hydroxyl radical addition to C5 of the pyrimidine, is produced via 350 nm photolysis of a 2,5-dimethoxyphenylsulfide precursor (2). Competition between O(2) and thiol for 1 suggests that the radical reacts relatively slowly with β-mercaptoethanol compared to other alkyl radicals. Overall, aryl sulfide 2 should be an effective precursor for the major hydroxyl radical adduct of thymidine in DNA.     

Action spectra for survival and spore photoproduct formation of Bacillus subtilis irradiated with short-wavelength (200-300 nm) UV at atmospheric pressure and in vacuo.

Spores of Bacillus subtilis are approximately ten times less likely to survive UV light irradiation in a vacuum than under atmospheric conditions. Photoproduct formation was studied in spores irradiated under ultrahigh vacuum (UHV) conditions and in spores irradiated at atmospheric pressure. In addition to the "spore photoproduct" 5-thyminyl-5,6-dihydrothymine (TDHT), which is produced in response to irradiation at atmospheric pressure, two additional photoproducts, known as the cis-syn and trans-syn isomers of thymine dimer, are produced on irradiation in vacuo. The spectral efficiencies for photoproduct formation in spores are reduced under vacuum conditions compared with atmospheric conditions by a factor of 2-6, depending on the wavelength. Because formation of TDHT does not increase after irradiation in vacuo, TDHT cannot be responsible for the observed vacuum effect. Vacuum specific photoproducts may cause a synergistic response of spores to the simultaneous action of UV light and UHV. An increased quantum efficiency, destruction of repair systems and formation of irreparable lesions are postulated for the enhanced sensitivity of B. subtilis spores to UV radiation in vacuo.     

Resonance-enhanced multiphoton ionisation photoelectron spectroscopy of the ClO radical: the C 2 sigma- state

A (2 + 1) one-colour resonance-enhanced multiphoton ionisation study is carried out on the C 2 sigma- state of the ClO radical in the one-photon energy range 29,500-31,250 cm-1. The ClO radical is produced by one-photon photolysis of ClO2 employing 359.2 nm photons derived from a separate laser. In this way a significant concentration of vibrationally excited ClO in its spin-orbit split X 2 pi omega (omega = 3/2 or 1/2) electronic ground state is produced. In addition to mass-resolved excitation spectra, kinetic-energy resolved photoelectron spectra for the X 3 sigma-(v+)<--C 2 sigma-(v' = 3-5) transitions are measured. These transitions are not completely Frank-Condon diagonal, and indicate a decrease in bond length on removal of the Rydberg electron from the C 2 sigma- state. In addition to an unambiguous assignment of the C 2 sigma- state, valuable information is obtained on the degree of vibrational excitation with which the nascent ClO radical is formed in the photolysis of ClO2. Analysis of the photoelectron spectra is supported by Franck-Condon calculations based on potential energy curves either from experimental spectroscopic parameters, or obtained by theoretical ab initio methods.     

Kinetic parameters and geometry of the transition state of acyl radical decarbonylation

Experimental data on acyl radical decomposition reactions (RC^·O → R^· + CO, where R = alkyl or aryl) are analyzed in terms of the intersecting parabolas method. Kinetic parameters characterizing these reactions are calculated. The transition state of methyl radical addition to CO at the C atoms is calculated using the DFT method. A semiempirical algorithm is constructed for calculating the transition state geometry for the decomposition of acyl radicals and for the reverse reactions of R^· addition to CO. Kinetic parameters (activation energy and rate constant) and geometry (interatomic distances in the transition state) are calculated for 18 decomposition reactions of structurally different acyl radicals. A linear correlation between the interatomic distance r ^#(C…C) (or r ^#(C…O)) in the transition state the enthalpy of the reaction (δH _e) is established for acyl decomposition reactions (at br _e = const). A comparative analysis of the enthalpies, activation energies, and interatomic distances in the transition state is carried out for the decomposition and formation of acyl, carboxyl, and formyl radicals.     

Atomic radical—molecule reactions F + CH<Subscript>3</Subscript>C≡CH: mechanistic study

The reaction of atomic radical F with propyne has been studied theoretically using ab initio quantum chemistry methods and transition state theory. The potential energy surface was calculated at the CCSD(T)/aug-cc-pVDZ (single-point) level using the UMP2/6-311++G(d,p) optimized structures. Two reaction mechanisms including the addition–isomerization–elimination reaction mechanism and the directed hydrogen abstraction reaction mechanism are considered. For the hydrogen abstraction reactions, i.e., the most probable evolution pathway in the title reaction, the HF formation occurs via direct abstraction mechanism dominantly and the H atom picked up by the atomic radical F should come mostly from the methyl group of normal propyne. On the other hand, for the addition–isomerization–elimination mechanism, the most feasible pathway should be the atomic radical F attacking on the C≡C triple bond in propyne (CH₃C≡CH) to form a weakly-bound adduct A1 with no barrier, followed by F addition to the C≡C triple bond to form the low-lying intermediate isomer 5. Subsequently, isomer 5 directly dissociates to P3 H₂CCCHF + H via transition state TS₅/P3. The other reaction pathways on the doublet PES are less competitive due to thermodynamical or kinetic factors. Furthermore, based on the analysis of the kinetics of all channels through which the addition and abstraction reaction proceed, we expect that the competitive power of reaction channels may vary with experimental conditions for the title reaction. The present work will provide useful information for understanding the processes of atomic radical F reaction with other unsaturated hydrocarbons. This material is available from author via E-mail.     

Independent generation of C5'-nucleosidyl radicals in thymidine and 2'-deoxyguanosine

The synthesis of the C5' tert-butyl ketone of thymidine 1a and 2'-deoxyguanosine 2 is achieved by reaction of 5'-C-cyano derivatives with tert-butyl lithium followed by acid hydrolysis. The 5'R configuration is assigned by X-ray crystal structure determination of an opportunely protected derivative of 1a. The (5'S)-isomers of both nucleosides are not stable, and a complete decomposition occurs in the reaction medium. The photochemistry of 1a and 2 effectively produced the thymidin-5'-yl radical and the 2'-deoxyguanosin-5'-yl radical, respectively. In the thymidine system, the C5' radical is fully quenched in the presence of a physiological concentration of thiols. In the 2'-deoxyguanosine system, the C5' radical undergoes intramolecular attack onto the C8-N7 double bond of guanine leading ultimately to the 5',8-cyclo-2'-deoxyguanosine derivative. The cyclization of the 2'-deoxyguanosin-5'-yl radical occurs with a rate constant of ca. 1x10(6) s-1 and is highly stereoselective affording only the (5'S)-diastereomer.     

Reactivity of 4‐thiothymidine with Fenton reagent investigated by UV‐visible spectroscopy and electrospray ionization mass spectrometry

The reactivity of the sulfur‐containing nucleoside 4‐thio‐(2′‐deoxy)‐thymidine usually abbreviated as 4‐thio‐thymidine, (S⁴‐TdR) under Fenton conditions, ie, in the presence of H₂O₂ and catalytic amounts of Fe(II), was investigated by UV‐vis spectroscopy and electrospray ionization single and tandem mass spectrometry (ESI‐MS and MS/MS). S⁴‐TdR hydroxylated on the S atom was found to be a key reaction intermediate, ultimately leading to (2′‐deoxy)‐thymidine usually abbreviated as thymidine, (TdR) as the main reaction product. This finding was in accordance with the outcome of the reaction between S⁴‐TdR and H₂O₂, previously investigated in our laboratory. On the other hand, the additional presence of ?OH radicals, induced by the Fe(II)/H₂O₂ combination, led to the increased generation of another interesting S⁴‐TdR product, already observed after its reaction with H₂O₂ alone, ie, the covalent dimer including a S? S bridge between two S⁴‐TdR molecules. More importantly, multihydroxylated derivatives of S⁴‐TdR and TdR were detected as peculiar products obtained under Fenton conditions. Among them, a product bearing an OH group both on the methyl group linked to the thymine ring and on the C5 atom of the ring was found to prevail. The results obtained during this study, integrated by those found previously in our laboratory, indicate 4‐thiothymidine as a promising molecular probe for the recognition, through a careful characterization of its reaction products, of the prevailing species among reactive oxygen species (ROS) corresponding to singlet‐state oxygen, hydrogen peroxide, and hydroxylic radical.     

Physical factors determining the activation energy of alkyl radical addition to unsaturated compounds

A parabolic model of the transition state is used for the analysis of experimental data (rate constants and activation energies) for reactions of addition of alkyl and phenyl radicals to multiple bonds of unsaturated compounds. The parameters describing the activation energy as a function of the enthalpy of the reactions were calculated from the experimental data. The activation energy depends also on the strength of the forming C−C bond, the presence of π-bonds in the α-position near the attacked C=C bond and the presence of polar groups in the monomer and radical. The empirical dependence of the activation energy of a thermoneutral addition reactionE _e0 on the dissociation energyD _e of the forming C−C bond was obtained:E _e0=(5.95±0.06)·10⁻⁴ D _e ² kJ mol⁻¹, indicating the important role of triplet repulsion in the formation of the transition state of radical addition. The contribution of the polar interaction to the activation energy of addition of polar radicals to polar monomers was calculated. Translated fromIzvestiya Akademii Nauk. Seriya Khimicheskaya, No. 3, pp. 445–450, March, 1999.     

Model Calculations of Radiation-Induced Damage in 1-Methyluracil:9-Ethyladenine

Detailed EPR and ENDOR experiments on the cocrystalline complex of 1-methyluracil:9-Ethyladenine (MUEA) have revealed that the major radiation-induced products observed at 10 K on MU are: MUEA1, a radical formed by net hydrogen abstraction from the N1-CH₃ methyl group, MUEA2, the MU radical anion, and MUEA3, the C5 H-addition radical. The following four products were observed on the adenine moiety at 10 K, MUEA4, the N3 protonated adenine anion, MUEA5, the native adenine cation, MUEA6, the amino deprotonated adenine cation, and MUEA7, the C8 H-addition radical formed by net H-addition to C8 of the adenine base. The geometries, energetics, and hyperfine properties of all possible radicals of MU and EA, the native anions and cations, as well as radicals formed via net hydrogen atom abstraction (deprotonated cations) or addition (protonated anions) were investigated theoretically. All systems were optimized using the hybrid Hartree–Fock–density functional theory functional B3LYP, in conjunction with the 6-31G(d,p) basis set of Pople and co-workers. Calculations of the anisotropic hyperfine couplings for all the radicals observed in MUEA are presented and are shown to compare favorably with the experimentally measured hyperfine couplings. The calculated ionizations potentials indicate that EA would be the preferred oxidation site. In MUEA, both the adenine cation and its N4-deprotonated derivative were observed. The calculated electron affinities indicate that MU would be the preferred reduction site. In MUEA radical, MUEA2 is a uracil reduction product, however the protonation state of this radical could not be determined experimentally. Calculations suggest that MUEA2 is actually the C4=O protonated anion.     

Kinetics and mechanism of the addition of the phthalimid-n-oxyl radical to the double bond of vinyl compounds

The kinetics of the decomposition of the phthalimid-N-oxyl radical (PINO) in acetic acid has been studied. The rate constants of the addition of the radical to T bonds of molecules of vinyl compounds – styrene, methyl methacrylate, acrylonitrile, and methyl acrylate – have been measured. It was shown that electron-donor substituents in the monomer molecule increase, while electron-acceptor substituents decrease the rate of addition. The reactivity of monomers in the elementary step of addition of the PINO radical decreases in the order CH₂=C(CH₃)C₆H₅ > CH₂=CHC₆H₅ > CH₂=C(CH₃)COOCH₃ > CH₂=CHCOOCH₃ > CH₂=CHCN.     

Self‐polyaddition of triethylsilyl perfluoroisopropenyl ether

The results on radical self‐polyaddition reactivity of two trialkylsilyl perfluoroisopropenyl ethers, triethysilyl perfluoroisopropenyl ether [CF₂?C(CF₃)OSi(C₂H₅)₃] (FTEE) and dimethylphenylsilyl perfluoroisopropenyl ether [CF₂?C(CF₃)OSi(CH₃)₂C₆H₅] (DMPE), and two perfluoroisopropenyl carboxylates, 2‐butyroxypentafluoropropene [CF₂?C(CF₃)OCOC₃H₇] (BuFPP) and 2‐(methoxyacetoxy)pentafluoropropene [CF₂?C(CF₃)OCOCH₂OCH₃] (MFPP), are described. Radical self‐polyaddition of FTEE afforded a polymer as high as 1.87 × 10⁴ in molecular weight in the presence of radical generators such as benzoyl peroxide and di‐tert‐butyl peroxide. DMPE gave only addition products with initiating radicals. BuFPP and MFPP scarcely yielded even addition products with radical. The mechanism that the self‐polyaddition of FTEE was initiated by the addition of radical onto the perfluoroisopropenyl group followed by a 1,5‐shift to afford a methyl radical that attacked the perfluoroisopropenyl group of another FTEE molecule is proposed. © 2003 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 41: 2743–2754, 2003     

Effect of glyoxylic oxime ether on radical polymerization of styrene

Methyl benzyloxyiminoacetate (MBOIA), a glyoxylic oxime ether, revealed different behaviors depending on the kinds of monomers used in the radical polymerization. MBOIA served as a retarder for styrene (St) and an inhibitor for vinyl acetate, whereas it showed little effect on the polymerization of methyl methacrylate. The retardation effect of MBOIA on the polymerization of St with dimethyl 2,2′‐azobisisobutyrate (MAIB) was examined in detail in benzene. The rate constant (k_x) of the reaction of MBOIA with polystyrene (PS) radical was 92 L/mol s at 50 °C, 112 L/mol s at 60 °C, and 143 L/mol s at 70 °C, indicating that the reactivity of MBOIA toward PS radical is less than that of St by a factor of about 3. The Arrhenius plot of k_x gave an activation energy of 20.3 kJ/mol. A nitrogen‐centered radical of a stationary state was observed by electron spin resonance (ESR) in the polymerization of St with MAIB at 60 °C in benzene in the presence of MBOIA, which is assignable to the radical (MBOIA ·) formed by addition of PS radical to MBOIA. The stationary MBOIA · concentration increased with increasing MBOIA concentration and then tended to be saturated at high concentrations. The rate constant of termination between MBOIA · radicals was 1.87 × l0⁵ L/mol s at 60 °C with ESR. © 2002 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 40: 2772–2781, 2002     

Additionen von SF5Cl und TeF5Cl an 〈CC〉 Doppelbindungen

Addition of SF₅Cl and TeF₅Cl on 〈C?C〉 double bonds Addition of SF₅Cl on 〈C?C〉 double bonds is investigated in a few examples. The results indicate a radical mechanism, in which the SF₅· free radical attacks the double bonds first. This is in agreement with many earlier findings. The direction of the addition is not changed by sterical influences. Sterically strained derivatives such as (SF₅)₂CH? CF₂Cl and SF₅(CF₃)₂C? CH₂Cl are obtained. In a single case the addition of TeF₅Cl on CH₂?CF₂ was possible, but the analogous reaction with SeF₅Cl was unsuccessful.     

Carboncarbon bond ligation between β-cyclodextrin and peptide by photo-irradiation

The formation of a stable covalent bond between β-cyclodextrin and different diphenyl-ketones (para-benzoylphenylalanine (pBz)Phe, para-benzoylbenzoic acid, pBzBz, and ketoprofen, KPF, under photo-irradiation is reported. These photo-probes have been incorporated into five peptides PEP1=Ac-Arg-Lys-Asp-Val-(pBz)Phe-NH₂; PEP2=Ac-Arg-Lys-Val-(pBz)Phe-NH₂; PEP3=Arg-Pro-(KPF)Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH₂; PEP4=N-(KPF)Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH₂; PEP5=Biot(O₂)-Apa-Arg-Arg-Pro-(pBzBz)Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met(O₂)-NH₂. The carboncarbon ligation occurs both in solution and in solid state and whatever the position of the diphenyl ketone, the ketyl radical is able to recombinate with β-cyclodextrin radical even in solid state. Photochemistry of diphenylketone is a complementary and orthogonal approach to chemical modifications of primary hydroxyl functions of β-cyclodextrin.     

Synthesis of polystyrene with high melting temperature through BDE/CuCl catalyzed polymerization

No matter what the polymerization manner was, polystyrene with unique highT _m (T _m = 170–285°C) was obtained through polymerization of styrene if the amount of BDE/CuCl catalyst was highly increased (mol ratio: St:CuCl = 25:1-2.5:1). Partial crystallinity of the PSt was observed by characterizations of X-ray diffraction and DSC. Spectra of¹H-NMR and¹³C-NMR showed that syndiotactic structure contained in the obtained PSt was 5% more than that in aPSt (atactic polystyrene). According to the proposed “coordinated radical cage” mechanism, the coordinated state between radical and catalyst center metal Cu should be more closely packed with increasing the BDE/CuCl catalyst amount, which was induced to partial stereospecific polymerization in the coordinated radical polymerization of St.