芳香氨基酸光敏化瞬态产物的光谱学及动力学表征

运用KrF激光闪光光解瞬态吸收光谱 ,以丙酮为光敏剂 ,研究了水溶液中芳香氨基酸的光化学反应 .通过动力学分析和猝灭实验 ,鉴别了光化学反应过程中的瞬态产物 ,获取了激发三重态的瞬态吸收光谱及动力学参数 .在丙酮存在下 ,色氨酸(Trp)和酪氨酸 (Tyr)的水溶液光解 ,分别观察到Trp激发三重态、N中心色氨酸自由基 (Trp/N·)和酪氨酸的酚氧自由基 (Tyr/O·) ,阐述了二者是丙酮三重态与Trp ,Tyr分别通过三重态 三重态 (T T)激发能转移和电子转移生成 ;苯丙氨酸 (Phe)不能与丙酮三重态进行激发能转移和电子转移 .进一步 ,在色氨酰酪氨酸 (Trp Tyr)敏化光解过程中 ,观察到分子内的电子转移 ,即Trp/N· Tyr→Trp Tyr/O·自由基的生成过程 .     

牛血清白蛋白的光损伤和光氧化机理

运用激光闪光光解瞬态吸收技术, 在266 nm激光激励下, 研究了牛血清白蛋白(BSA)光损伤和被SO4-·单电子氧化的反应机理, 表征了反应过程中生成的自由基. 结果表明, 在266 nm激光照射下, BSA可同时发生光电离和光激发, 生成色氨酸阳离子自由基(Trp/NH+·), 由Trp/NH+·快速脱质子形成的色氨酸中性自由基(Trp/N·)及色氨酸三重激发态(3Trp*), 3Trp*再与酪氨酸(Tyr)发生分子内电子转移生成酪氨酸中性自由基(Tyr/O·). 在SO4-·单电子氧化的反应中, 借助减谱技术, 求得BSA中Tyr和色氨酸(Trp)自由基的表观生成速率常数, 但未发现分子内电子转移现象, 阐明了SO4-·自由基是通过与BSA中的Tyr和Trp发生电子转移反应来氧化BSA的, SO4-·氧化BSA的反应速率常数为1.51×10¹⁰ L·mol^-1·s^-1, 从而为进一步研究血清白蛋白的氧化还原代谢过程提供理论基础.     

烷基-全氟酰氧基氮氧自由基——全氟酰基过氧化物与脂肪族伯、仲胺单电子转移反应的EPR研究

EPR研究表明,全氟酰基过氧化物在室温下可将脂肪族仲胺氧化成相应的稳定双烷基氮氧自由基。它们与RNH₂的氧化反应是单电子转移过程,生成烷基-全氟酰氧基氮氧自由基RN(O^·)OCOR_F;当R为叔烷基时,还生成RN(O^·)R、RN(O^·)H、RN(O^·)R_F和RN(O^·)R_F′(R_F′=R_F-CF₂)等4种稳定性差别很大的氮氧自由基。EPR研究结果揭示了该反应机理的重要信息。     

2-蒽醌磺酸三重态与高聚鸟嘌呤核苷酸相互作用的激光光解

利用248 nm(KrF)激光光解技术研究了乙腈-水(97︰3)溶液中2-蒽醌磺酸激发三重态电子转移光氧化高聚鸟嘌呤核苷酸的原初过程.直接检测了上述电子转移氧化反应生成的阴阳离子自由基对的瞬态吸收光谱,分别获取了各自的表观反应速率常数,协同反应的自由能变化,阐明了电子转移反应的三重态机理.     

色氨酸及其肽对cis-syn型1，3-二甲基胸腺嘧啶二聚体的光敏化裂解作用

通过荧光猝灭实验和测定二聚体裂解程度的辐照实验，研究色氨酸（Trp）及 其二肽色氨酰苯丙氨酸（Trp-Phe）对cis-syn型1，3-二甲基胸腺嘧啶二聚体（ DMTD）的光敏化裂解作用。结果表明，色氨酸及其二肽在较强光（λ > 290 nm） 辐照下，主要通过双光子电离生成的光合电子（e_(aq)~-）导致二聚体裂解，其次 ，通过激发单重态与二聚体间的电子转移导致二聚体裂解。另外一导致二聚体裂解 的可能途径：色氨酸残基激发三重态与二聚体间的电子转移光敏化二聚体裂解。     

金属酞菁激发态与氧的相互作用

比较了几种金属酞菁光敏产生单重态氧和超氧负离子的能力,结果表明它们产生¹O₂的能力与中心金属的电子结构有关,取决于三重态寿命和量子产率。顺序如下:Zn>Ga>Cu>H₂>Al>Co。产生O₂^·-的能力不仅与三重态寿命和量子产率有关,也与激发能和氧化还原电位有关。其顺序如下:Ga>Al>Cu>Zn。还研究了酪氨酸与镓酞菁激发态相互作用,酪氨酸猝灭镓酞菁荧光。在除氧条件光激发下,酪氨酸猝灭镓酞菁的激发三重态发生电子转移,检测到GaTSPc^-在560nm处的瞬态吸收,在氧的存在下进一步反应生成O₂^·-。     

维生素K3的激发三重态与色氨酸、酪氨酸电子转移氧化反应的激光闪光光解研究

利用时间分辨的激光闪光光解技术研究了乙腈-水混合溶液(1:1,V/V)中2-甲基萘醌(通常称为维生素K3)的激发三重态对色氨酸、酪氨酸的光敏氧化机理.通过瞬态吸收光谱的变化可以推断维生素K3的激发三重态可以与色氨酸、酪氨酸发生电子转移反应,反应形成的维生素K3阴离子自由基的吸收峰可以直接从瞬态吸收谱图中观察到.维生素K3与色氨酸、酪氨酸的电子转移反应的速率分别为1.1×109和0.6×109L·mol-1·s-1.吉布斯自由能(ΔG)的计算结果表明维生素K3的激发三重态与色氨酸、酪氨酸电子转移反应在热力学上是可行的.     

稀土金属离子及pH诱导牛血清白蛋白构象变化的同步荧光光谱研究

用同步荧光法并辅以普通荧光法对不同浓度稀土离子及pH诱导的牛血清白蛋白(BSA)构象变化进行了详细研究, 发现稀土离子使色氨酸(Trp)残基的荧光蓝移, 荧光强度降低; 酪氨酸(Tyr)残基荧光峰位置不变, 稀土离子浓度较低时, 荧光峰强度降低, 而当浓度较高时, Tyr残基荧光峰强度反而增强. 据此推断了稀土离子与BSA结合反应中Trp残基微环境和Tyr残基微环境及构象的变化并与pH引起的变化进行了比较.     

4-硫代尿嘧啶与色氨酸光致电子转移反应研究

RNA与蛋白质相互作用是生物体进行生命活动的基础,光活化核苷酸引发的共价交联是研究其相互作用的有效手段.对其机理的研究有助于理解并调控交联的位点及氨基酸,因此探测关键的中间体来揭示机理很有必要.本工作选择光活化核苷酸4-硫代尿嘧啶（4-TU）和色氨酸（TrpH）为模型体系,通过激光闪光光解技术探测瞬态中间体并由此揭示机理.实验发现4-TU三重态与TrpH发生电子转移反应,瞬态吸收光谱上观察到4-TU^·-及TrpH^·+,以及TrpH^·+脱质子生成的Trp^·.通过测量4-TU三重态衰减动力学得到电子转移反应的速率常数为2.88×10⁹ L·mol^-1·s^-1,并研究了pH值对反应的影响.电子转移驱动力ΔG为-0.15 eV,是能量有利的过程.结果表明电子转移反应是光致共价交联过程中关键的第一步反应,引发了后续的质子转移及自由基交联过程.     

核黄素(维生素B2)的光物理和光化学性质

应用时间分辨的激光光解吸收技术, 详细研究了核黄素(维生素B2)的光物理和光化学性质. 在337 nm激光作用下, 可产生激发三重态(3RF*); 而在248 nm激光作用下, 既可产生激发三重态, 又可光电离生成水合电子和阳离子自由基(). 探索了激发三重态的各种反应性(能量转移、 氧化还原和抽氢反应等)及其潜在的意义, 并用自由基氧化核黄素产生阳离子自由基, 进一步验证了光解结果.     

Characterization of transient species in laser photolysis of aromatic amino acids using acetone as photosensitizer

The photochemical processes of aromatic amino acids were investigated in aqueous solution using acetone as photosensitizer by KrF (248 nm) laser flash photolysis. Laser-induced transient species were characterized according to kinetic analysis and quenching experiments. The intermediates recorded were assigned to the excited triplet state of tryptophan, the radicals of tryptophan and tyrosine. The excited triplet state of tryptophan produced via a triplet-triplet excitation transfer and the radicals arising from electron transfer reaction has been identified. Neither electron transfer nor energy transfer between triplet acetone and phenylalanine can occur in photolysis of phenylalanine aqueous solution which contains acetone. Furthermore, triplet acetone-induced radical transformation: Trp/N-Tyr→Trp-Tyr/O was observed directly in photolysis of dipeptide (Trp-Tyr) aqueous solution containing acetone, and the transformation resulting from intramolecular electron transfer was suggested.     

Photoinduced electron transfer reactions of rose bengal and selected electron donors

The photoinduced electron transfer reactions of the triplet state of rose bengal (RB) and several electron donors were investigated by the complementary techniques of steady state and time-resolved electron paramagnetic resonance (EPR) and laser flash photolysis (LFP). The yield of radicals varied with the light fluence rate, RB concentration and, in particular, the electron donor used. Thus for L-dopa (dopa, dihydroxyphenylalanine) only 10% of RB anion radical (RB√−) was produced, with double the yield observed with NADH (NAD, nicotinamide adenine dinucleotide) as quencher and more than three times the yield observed with ascorbate as quencher. Quenching of the RB triplet was both reactive and physical with total quenching rate constants of 4 × 10⁸ mol⁻¹ dm³ s⁻¹ and 8.5 × 10⁸ mol⁻¹ dm³ s⁻¹ for ascorbate and NADH respectively. The rate constant for the photoinduced electron transfer from ascorbate to RB triplet was 1.4 × 10⁸ mol⁻¹ dm³ s⁻¹ as determined by Fourier transform EPR (FT EPR). FT EPR spectra were spin polarized in emission at early times indicating a radical pair mechanism for the chemically induced dynamic electron polarization. Subsequent to the initial electron transfer production of radicals, a complex series of reactions was observed, which were dominated by processes such as recombination, disproportionation and secondary (bleaching) reactions.

It was observed that back electron transfer reactions could be prevented by mild oxidants such as ferric compounds and duroquinone, which were efficiently reduced by RB√−.     

A laser flash photolysis study of amino acids and dipeptides using 4-nitroquinoline 1-oxide as a photosensitizer: The pH dependence

The pH effects on the photochemical reaction of amino acids and related dipeptides with 4-nitroquinoline 1-oxide (4NQO) as a photosensitizer have been investigated by laser flash photolysis. The obtained kinetic parameters show that the electron transfer from Tryptophan (Trp), Tyrosine (Tyr) as well as dipeptides containing Trp and/or Tyr residue to triplet 4NQO (^T4NQO) are efficient, but inefficient from methionine (Met) and dipeptides containing neither Trp nor Tyr. The result was supported by the calculated values of the free energy change from measured oxidation potentials for the electron transfer. It was demonstrated that Trp and Tyr residues are initial reaction sites with ^T4NQO, while Tyr/O^? radical may be final species for Trp-Tyr dipeptide. In acidic aqueous solutions, the self-quenching rate constants of ^T4NQO and the rate constants of electron transfer from amino acids to ^T4NQO decrease with decreasing pH. In alkaline solutions, amino acids are easily oxidized by 4NQO under irradiation of laser pulse, and no transient absorption signal was observed.     

分子光化学贮存太阳能 Ⅲ.双环[2,2,1]-2,5-庚二烯的光诱导电子转移敏化异构化反应

以N,N,N′,N′-四甲基联苯二胺、2,6-二甲氧基萘和2,7-二甲氧基萘为光敏剂,在正己烷溶液中实现了双环[2,2,1]-2,5-庚二烯到四环[2,2,1,0^2,6,0^3,5]庚烷的异构化。测定了反应的量子产率。讨论了反应机理。通过激发态的光敏剂与二烯之间的电子转移反应,形成单重态和三重态处于平衡状态的离子自由基对中间体。处于溶剂笼中的三重态离子自由基对经电子反传,产生激发三重态二烯。最后该激发态二烯经分子内[2+2]环合加成反应异构化为四环烷。     

Cation-π effects in the complexation of Na+ and K+ with Phe, Tyr, and Trp in the gas phase

Na⁺ and K⁺ gas-phase affinities of the three aromatic amino acids Phe, Tyr, and Trp were measured by the kinetic method. Na⁺ binds these amino acids much more strongly than K⁺, and for both metal ions the binding strength was found to follow the order Phe ≤ Tyr < Trp. Quantum chemical calculations by density functional theory (DFT) gave the same qualitative ordering, but suggested a somewhat larger Phe/Trp increment. These results are in acceptable agreement with predictions based on the binding of Na⁺ and K⁺ to the side chain model molecules benzene, phenol, and indole, and are also in reasonable agreement with the predictions from purely electrostatic calculations of the side-chain binding effects. The binding energies were compared with those to the aliphatic amino acids glycine and alanine. Binding to the aromatic amino acids was found to be stronger both experimentally and computationally, but the DFT calculations indicate substantially larger increments relative to alanine than shown by the experiments. Possible reasons for this difference are discussed. The metal ion binding energies show the same trends as the proton affinities.     

Dissociations of copper(II)-containing complexes of aromatic amino acids: radical cations of tryptophan, tyrosine, and phenylalanine

The dissociations of two types of copper(II)-containing complexes of tryptophan (Trp), tyrosine (Tyr), or phenylalanine (Phe) are described. The first type is the bis-amino acid complex, [Cu(II)(M)(2)].(2+), where M = Trp, Tyr, or Phe; the second [Cu(II)(4Cl-tpy)(M)].(2+), where 4Cl-tpy is the tridendate ligand 4'-chloro-2,2':6',2'-terpyridine. Dissociations of the Cu(ii) bis-amino acid complexes produce abundant radical cation of the amino acid, M.(+), and/or its secondary products. By contrast, dissociations of the 4Cl-tpy-bearing ternary complexes give abundant M.(+) only for Trp. Density functional theory (DFT) calculations show that for Tyr and Phe, amino-acid displacement reactions by H(2)O and CH(3)OH (giving [Cu(II)(4Cl-tpy)(H(2)O)].(2+) and [Cu(II)(4Cl-tpy)(CH(3)OH)].(2+)) are energetically more favorable than dissociative electron transfer (giving M.(+) and [Cu(I)(4Cl-tpy)](+)). The fragmentation pathway common to all these [Cu(II)(4Cl-tpy)(M)].(2+) ions is the loss of NH(3). DFT calculations show that the loss of NH(3) proceeds via a "phenonium-type" intermediate. Dissociative electron transfer in [Cu(II)(4Cl-tpy)(M-NH(3))].(2+) results in [M-NH(3)].(+). The [Phe-NH(3)] (+) ion dissociates facilely by eliminating CO(2) and giving a metastable phenonium-type ion that rearranges readily into the styrene radical cation.     

ENERGY CONVERSION IN THE DECAY OF TRIPLET LUMIFLAVIN IN THE PRESENCE OF FERRI- AND FERROCYANIDE

Abstract— Flash photolysis at 450 nm has been used to study the quenching of the excited triplet state of lumiflavin and the transient species formed in subsequent reactions in deaerated phosphate buffer (pH 6.9).
The effect of the presence of ferricyanide on the life time of triplet lumiflavin has been studied. The results suggest an energy transfer reaction without concurrent electron transfer reactions. The rate constant for the process was 2.8 times 10⁹ M ^-1 s^-1. The analogous reaction with ferrocyanide could not be observed because of the efficient electron transfer reaction (δG = -20.6 kcal mol^-1) leading to the formation of the semireduced lumiflavin and ferricyanide. The rate constant for this reaction was 3.3 times 10⁹ M ^-1 s^-1. The semireduced lumiflavin radical was found to disappear in a second order reaction with a rate constant of 1.7 times 10⁹ M ^-1 s^-1. It was found to react with ferricyanide with a rate constant of 0.7 times 10⁹ M ^-1 s^-1.
A model for the various photochemical and photophysical processes involved in the decay and quenching of the lumiflavin triplet state is suggested and discussed.     

ALKALINE QUENCHING OF BACTERIORHODOPSIN TRYPTOPHANYL FLUORESCENCE: EVIDENCE FOR AQUEOUS ACCESSIBILITY OR A HYDROGEN-BONDED CHAIN

Abstract— Comparison between Trp fluorescence yields of membrane-bound bacteriorhodopsin (BR) and retinylidene-free bacterioopsin (BO) is consistent with a model in which all eight Trp residues are active fiuorophores in the latter, while the emission of all but two residues in the former is lost by energy transfer to retinal. The visible chromophore of BR is progressively bleached with increasing pH. Up to pH 12 this bleaching is reversed on reneutralization; but above this the change is irreversible with the appearance of a new absorption band characteristic of free retinal. Emission yields for both proteins decrease with increasingly alkaline pH in a manner typical of energy transfer to weakly-fluorescent tyrosinate. The limiting yields, reached at a pH corresponding to that producing irreversible bleaching of the visible chromophore, agree with an integral value of one remaining active Trp fluorophore in BR and between one and two in BO and show that the bulk of Trp are within the 11 Å Förster energy-transfer distance of Tyr accessible to OH⁻. Current models of the native protein structure of BR arrange the polypeptide chain primarily in a bundle of seven helical segments with axes perpendicular to the lipid bilayer plane and with buried polar residues, including Trp and Tyr, located at intrahelical surfaces. An interpretation of the observed accessibility of buried Tyr to OH⁻ is that an aqueous region exists within the protein structure. Moreover, this putative aqueous region must be close to the retinylidene chromophore and thus may be associated with the light-driven ion transport system. The results are also compatible with energy transfer to internal Tyr residues which are connected via a chain of phenolate hydrogen bonds to a surface Tyr.     

香豆素酮类化合物的瞬态研究——Ⅲ.香豆素酮和碘鎓盐间的电子转移

本工作采用激光闪光光解法对香豆素酮类化合物敏化碘鎓盐化合物问题进行了研究。结果表明:碘鎓盐化合物能通过电子转移机理猝灭香豆素酮的激发三重态。工作中还用甲基紫精(PQ²⁺)为模型化物,观察到它也能使香豆素酮的三重态猝灭,同时可看到在位于610nm处的PQ⁺生成。这些结果说明,在发生电子转移的反应中香豆素酮是电子给体,按Weller公式的计算结果也表明它们之间可发生电子转移反应。