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The carbonate radical (CO 3 (*-)) is of importance in biology and chemistry. We used pulse radiolysis to generate the CO 3 (*-) radical and show there is no reaction with oxygen. However, in the presence of ammonia the CO 3 (*-) radical is removed by NO (*), which itself arises from the scavenging of NH 2 (*) by oxygen, and the mechanism of this process is reported. The CO 3 (*-) radical shows complex decay patterns in the presence of ammonia, which can be understood as a balance between the radical-radical reaction CO 3 (*-) + CO 3 (*-) and CO 3 (*-) + NH 2 (*) (the amino radical). Also, we report reactivity with glycine and alanine and with melanin models. The CO 3 (*-) reacts with both dopa-melanin (DM, a model of black eumelanin) and with cysteinyl-dopa-melanin (CDM, a model of red/blond phaeomelanin). However, the reaction rate constant is much higher with CDM than with DM.

Formula chem. The chemical and spectroscopic characterization of 5,6-indolequinones and their semiquinones, key transient intermediates in the oxidative conversion of 5,6-dihydroxyindoles to eumelanin biopolymers, is a most challenging task. In the present paper, we report the characterization of a novel, relatively long-lived 5,6-indolequinone along with its semiquinone using an integrated chemical, pulse radiolytic, and computational approach. The quinone was obtained by oxidation of 5,6 -dihydroxy-3-iodoindole (la) with o-chloranil in cold ethyl acetate or aqueous buffer: it displayed electronic absorption bands around 400 and 600 nm, was reduced to la with Na 2 S 2 O 4 , and reacted with o-phenylenediamine to give small amounts of 3-iodo- 1H -pyrrolo[2,3-b ]phenazine (2). The semiquinone exhibited absorption maxima at 380 nm (sh) and 520 nm and was detected as the initial species produced by pulse radiolytic oxidation of la at pH 7.0. DFT investigations indicated the 6-phenoxyl radical and the N-protonated radical anion as the most stable tautomers for the neutral and anion forms of the semiquinone, respectively. Calculated absorption spectra in water gave bands at 350 (sh) and 500 nm for the neutral form and at 310 and 360 (sh) nm for the anion. Disproportionation of the semiquinone with fast second-order kinetics (2k = 1.1 x 10 10 M -1 s -1 ) gave a chromophore with absorption bands resembling those of chemically generated la quinone. Computational analysis predicted la quinone to exist in vacuo as the quinone-methide tautomer, displaying low energy transitions at 380 and 710 nm, and in water as the o-quinone, with calculated absorption bands around 400 and 820 nm. A strong participation of a p orbital on the iodine atom in the 360-380 nm electronic transitions of the o-quinone and quinone-methide was highlighted. The satisfactory agreement between computational and experimental electronic absorption data would suggest partitioning of la quinone between the o-quinone and quinone-methide tautomers depending on the medium.

Eumelanin structure and build up is still a challenge. Two models have been proposed to account for the unique properties of eumelanin: one put forward by Simon (Clancy and Simon, 2001) envisages a planar oligomer of 5,6-dihydroxyindole (DHI) /5,6-dihydroxyindole-2-carboxylic acid (DHICA) units which assembles through -stacking and side-on interactions, while the other (Cheng et al, 1994) depicts five to seven monomers of 5,6-indolequinone arranged in a plane, forming 3-4 layers stacked 3.4 Å apart. To probe either of these hypotheses, we underwent a systematic characterisation of oligomers of DHI and DHICA and their oxidation chemistry. Here we report the use of combined pulse radiolysis and quantum mechanical calculations to inquire into the nature of the species formed by oxidation of three dimers of DHI, namely 5,6,5’,6’-tetrahydroxy-2,2’-biindolyl, 5,6,5’,6’-tetrahydroxy-2,4’-biindolyl and 5,6,5’,6’-tetrahydroxy-2,7’-biindolyl. Pulse radiolytic oxidation of 2,4’- and 2,7’-biindolyls led to semiquinones absorbing around 450 nm, which decayed to give the corresponding quinones (500-550 nm). 2,2’-Biindolyl, on the other hand, furnished a semiquinone (max 480 nm) which disproportionated to give a relatively stable quinone ( max 580 nm). A quantum mechanical investigation of o-quinone, quinone imine and quinone methide structures of the dimers suggested that in the oxidised forms they exist mainly as 2-substituted extended quinomethane tautomers stabilised by intramolecular hydrogen bonds, reaching a planar conformation which is not favoured for the reduced forms. Overall, these results indicate that oxidation of oligomers to quinonoid species is critical to achieve planar assemblies which can interact by pi-stacking

Investigation of the oxidative polymerization of 5,6-dihydroxyindoles remains one of the most fruitful approaches toward definition of the structure of eumelanins. Previous studies by our group led to isolation of a collection of dimers and trimers which yielded the first insight into the stages of melanogenesis beyond dopachrome rearrangement. Attempts to isolate and characterize higher oligomeric intermediates were thwarted by the marked complexity of the product patterns. Recently, we reported an alternative approach to this issue involving biomimetic oxidation of 5,6-dihydroxyindole dimers, leading for the first time to novel tetramer species. For example, oxidation of the 4,4’dimer of 5,6-dihydroxyindole-2-carboxylic acid (DHICA) yielded a mixture of isomeric tetramers linked through the 7-position and featuring chiroptical properties arising from atropoisomerism at the biphenyl linkages. Herein, we report the pulse radiolytic investigation of the oxidation of 2,4’ and 2,7’dimers of 5,6-dihydroxyindole (DHI). The results provided evidence for the formation of two different semiquinone species for each dimer, which decayed with second order kinetics to quinonoid product(s) exhibiting absorption maxima at 530 and 550 nm. The symmetric 2,2’dimer on the other hand furnished a single semiquinone disproportionating at a much slower rate to give a quinone with absorption maximum at 580 nm. Peroxidase/H2O2 oxidation of the 2,4’ and 2,7’ dimers of DHI gave tetramer products, the major of which were characterized by extensive 2D-NMR and mass spectrometry. The results disclosed unexpected differences in the mode of coupling of the dimers compared to DHI

1,2-Dehydro-N-acetyldopamine (dehydro-NADA) is an important catecholamine derivative involved in the cross-linking of insect cuticular components during sclerotization. Since sclerotization is a vital process for the survival of insects, and is closely related to melanogenesis, it is of interest to unravel the chemical mechanisms participating in this process. The present paper reports on the mechanism by which dehydro-NADA is oxidatively activated to form reactive intermediate(s) as revealed by pulse radiolysis, electron spin resonance spectroscopy, high performance liquid chromatography, and ultraviolet-visible spectroscopic analysis. Pulse radiolytic one-electron oxidation of dehydro-NADA by N3. (k = 5.3 x 10(9) M-1 s-1) or Br2.- (k = 7.5 x 10(8) M-1 s-1) at pH6 resulted in the rapid generation of the corresponding semiquinone radical, lambda max 400 nm, epsilon = 20,700 M-1 cm-1. This semiquinone decayed to form a second transient intermediate, lambda max 485 nm, epsilon = 8000 M-1 cm-1, via a second order disproportionation process, k = 6.2 x 10(8) M-1 s-1. At pH 6 in the presence of azide, the first order decay of this second intermediate occurred over milliseconds; the rate decreases at higher pH. At pH 6 in the presence of bromide, the intermediate decayed much more slowly over seconds, k = 0.15 s-1. Under such conditions, the dependence of the first order decay constant upon parent dehydro-NADA concentration led to a second order rate constant of 8.5 x 10(2) M-1 s-1 for reaction of the intermediate with the parent, probably to form benzodioxan "dimers." (The term dimer is used for convenience; the products are strictly bisdehydrodimers of dehydro-NADA (see "Discussion" and Fig. 11)) Rate constants of 5.9 x 10(5), 4.5 x 10(5), 2.8 x 10(4) and 3.5 x 10(4) M-1 s-1 were also obtained for decay of the second intermediate in the presence of cysteine, cysteamine, o-phenylenediamine, and p-aminophenol, respectively. By comparison with the UV-visible spectroscopic properties of the two-electron oxidized species derived from dehydro-NADA and from 1,2-dehydro-N-acetyldopa methyl ester, it is concluded that the transient intermediate exhibiting absorbance at 485 nm is the quinone methide tautomer of the o-quinone of dehydro-NADA. Sclerotization of insect cuticle is discussed in the light of these findings.

We report the direct observation of the quenching of the weakly absorbing transient due to the amino radical by oxygen and, hence determine, by a totally direct method, the corresponding rate constant (k = (1.1 +/- 0.1) x 10(9) dm3 mol(-1) s(-1)). We also report the rate constants for the reactions of the amino radical with several amino acids and models of black eumelanin and blond/red phaeomelanin. These reactions lead to a mechanism, based on free radicals, that can explain why ammonia is useful in commercial hair (melanin) bleaching, avoiding excessive amino acid (hair protein) damage.

The particular chromophoric structure of C(70)Ph(10), which consists of two cage-centered π-electron systems, makes its photophysical properties an exception to those found for other phenylated [70]fullerenes C(70)Ph(2n) (n=2-4). For these other C(70)Ph(2n) species, their intrinsic photophysical properties undergo smooth transitions as a function of n.

Using pulse radiolysis and laser flash photolysis, we have investigated the reactions of the deleterious species, e(-)(aq), HOz.rad;, O(2)(*)(-) and O(2)((1)Delta(g)) with 10 water-soluble cyclopropyl-fused C(60) derivatives including a mono-adduct dendro[60]fullerene (d) and C(60) derivatives based on C(60)[C(COOH)(2)](n=2-6), some of which are known to be neuroprotective in vivo. The rate constants for reactions of e(-)(aq) and HOz.rad; lie in the range 0.5-3.3 x 10(10) M(-1) s(-1). The d and bis-adduct monoanion radicals display sharp absorption peaks around 1000 nm (epsilon = 7 000-11 500 M(-1) cm(-1)); the anions of the tris-, tetra-, and penta-adduct derivatives have broader, weaker absorptions. The monohydroxylated radicals have their most intense absorption maxima around 390-440 nm (epsilon = 1000-3000 M(-1) cm(-1)). The anion and hydroxylated radical absorption spectra display a blue-shift as the number of addends increases. The radical anions react with oxygen (k approximately 10(7)-10(9) M(-1) s(-1)). The reaction of O(2)(*)(-) with the C(60) derivatives does not occur via an electron transfer. The rate constants for singlet oxygen reaction with the dendrofullerene and eee-derivative in D(2)O at pH 7.4 are k approximately 7 x 10(7) and approximately 2 x 10(7) M(-1) s(-1) respectively, in contrast to approximately 1.2 x 10(5) M(-1) s(-1) for the reaction with C(60) in C(6)D(6). The large acceleration of the rates for electron reduction and singlet oxygen reactions in water is due to a solvophobic process.

Eumelanogenesis and phaeomelanogenesis diverge at an early stage in pigment formation, namely at the point where dopaquinone, the initial product of tyrosine oxidation by tyrosinase, undergoes one of two types of reaction: either (1) a reductive endocyclisation in which a Michael addition of the side-chain amino group takes place; or (2) a reductive addition of cysteine to give cysteinyldopa. In the former case, the product cyclodopa, is known rapidly to undergo a redox exchange reaction with dopaquinone to yield dopachrome, the precursor of the eumelanogenic pathway. In the second instance, cysteinyldopa is regarded as leading to the formation of benzothiazoles, which are characteristic of phaeomelanin. The precursor molecule of the phaeomelanic pathway is cysteinyldopaquinone. We have examined quantitatively the role of dopaquinone in the non-enzymatic oxidation of 5-S-cysteinyldopa using pulse radiolysis and have demonstrated that the redox exchange reaction between dopaquinone and 5-S-cysteinyldopa occurs spontaneously with a rate constant of 8.8 x 10(5) M(-1) sec(-1). This study has also enabled an improved estimate ofor = 4 x 10(7) M(-1) sec(-1) to be obtained for the rate constant of the reaction of dopaquinone with cyclodopa. Calculations utilising these figures and estimates of the rate constants for the other reactions in early melanogenesis, demonstrate that, whilst similar pathways are invoked, the phaeomelanic pathway predominates in the presence of cysteine, irrespective of the availability of dopaquinone and thus independently of the rate of tyrosinase-catalysed oxidation. This suggests that the balance between the formation of eumelanin and phaeomelanin is regulated principally by the availability of cysteine at the site of melanogenesis.

The human skin and eye melanin is commonly viewed as an efficient photoprotective agent. To elucidate the molecular mechanism of the melanin-dependent photoprotection, we studied the interaction of two synthetic melanins, dopa-melanin and cysteinyldopa-melanin, with a wide range of oxidising and reducing free radicals using the pulse radiolysis technique. We have found that although both types of free radicals could efficiently interact with the synthetic melanins, their radical scavenging properties depended, in a complex way, on the redox potential, the electric charge and the lifetime of the radicals. Repetitive pulsing experiments, in which the free radicals, probing the polymer redox sites, were generated from four different viologens, indicated that the eumelanin model had more reduced than oxidised groups accessible to reaction with the radicals. Although with many radicals studied, melanin interacted via simple one-electron transfer processes, the reaction of both melanins with the strongly oxidising peroxyl radical from carbon tetrachloride, involved radical addition. Our study suggests that the free radical scavenging properties of melanin may be important in the protection of melanotic cells against free radical damage, particularly if the reactive radicals are generated in close proximity to the pigment granules.

Rate constants have been determined for reactions between biologically significant thiols, represented by cysteine and glutathione, and a series of 10 4-substituted o-quinones, and unsubstituted o-quinone itself, generated by rapid disproportionation of the semiquinones formed from the corresponding catechols by pulse radiolysis. The quantitative structure-reactivity relationships were investigated by examining the correlation between the rate constants and various Hammett and other parameters characterizing the electronic nature of the substituents. From these relationships, it can be concluded that the o-quinone reactivity with thiols increases with the electron-withdrawing capacity of the substituent groups and that this effect is principally due to resonance effects. Such relationships allow the prediction of likely reactivities with cellular thiols of further o-quinones whose 4-substituents have known electronic parameters. These reactivities are likely to be one of the critical factors determining overall cytotoxicity, assisting in the choice of improved melanogenesis-targeted anti-melanoma drugs.

1,2-Dehydro-N-acetyldopamine (dehydro-NADA) is an important catecholamine derivative involved in the cross-linking of insect cuticular components during sclerotization. Since sclerotization is a vital process for the survival of insects, and is closely related to melanogenesis, it is of interest to unravel the chemical mechanisms participating in this process. The present paper reports on the mechanism by which dehydro-NADA is oxidatively activated to form reactive intermediate(s) as revealed by pulse radiolysis, electron spin resonance spectroscopy, high performance liquid chromatography, and ultraviolet-visible spectroscopic analysis. Pulse radiolytic one-electron oxidation of dehydro-NADA by N3. (k = 5.3 x 10(9) M-1 s-1) or Br2.- (k = 7.5 x 10(8) M-1 s-1) at pH6 resulted in the rapid generation of the corresponding semiquinone radical, lambda max 400 nm, epsilon = 20,700 M-1 cm-1. This semiquinone decayed to form a second transient intermediate, lambda max 485 nm, epsilon = 8000 M-1 cm-1, via a second order disproportionation process, k = 6.2 x 10(8) M-1 s-1. At pH 6 in the presence of azide, the first order decay of this second intermediate occurred over milliseconds; the rate decreases at higher pH. At pH 6 in the presence of bromide, the intermediate decayed much more slowly over seconds, k = 0.15 s-1. Under such conditions, the dependence of the first order decay constant upon parent dehydro-NADA concentration led to a second order rate constant of 8.5 x 10(2) M-1 s-1 for reaction of the intermediate with the parent, probably to form benzodioxan "dimers." (The term dimer is used for convenience; the products are strictly bisdehydrodimers of dehydro-NADA (see "Discussion" and Fig. 11)) Rate constants of 5.9 x 10(5), 4.5 x 10(5), 2.8 x 10(4) and 3.5 x 10(4) M-1 s-1 were also obtained for decay of the second intermediate in the presence of cysteine, cysteamine, o-phenylenediamine, and p-aminophenol, respectively. By comparison with the UV-visible spectroscopic properties of the two-electron oxidized species derived from dehydro-NADA and from 1,2-dehydro-N-acetyldopa methyl ester, it is concluded that the transient intermediate exhibiting absorbance at 485 nm is the quinone methide tautomer of the o-quinone of dehydro-NADA. Sclerotization of insect cuticle is discussed in the light of these findings.

The various effects of ionising radiation in biological systems are generally thought to be a consequence of chemical modifications induced within DNA (Painter, 1979). Indeed DNA is generally accepted to be the major target which is sensitive to ionising radiation resulting in the various biological effects determined. Following the initial energy deposition events, the radiation-induced damages to DNA most generally measured in biological systems are single and double strand breaks and modified bases. At present, there are little experimental data on the time course of the early processes which occur following the initial energy deposition events (~10–15 s). The present biophysical models (Goodhead, this volume) generally relate the distribution of energy deposition events to the biological effects of radiation quality.

SummaryUsing diffuse reflectance pulse radiolysis, it has been demonstrated from spectral characteristics of the resulting transients that the chemical events following irradiation of DNA depend upon its state of hydration.

Numerous improvements have been made to the Paterson Institute linear accelerator since its installation in 1967. New light sources, improved light guidance, smaller cells and a wider range of photo-detecting devices are now in routine use. Data are collected and processed by a computer-based method which has replaced the original oscilloscope-based system. Processes taking place over more than a few seconds can be studied with an arrangement combining pulse radiolysis with an ordinary spectrophotometer and arrangements for “single-shot” studies of faster processes are now being designed. Detection methods are also available which do not rely on transmission of light, and transient changes in conductivity can be measured. Among the systems which have been extensively studied are the Fricke dosimeter, in which measured overall yields can now be quantitatively correlated with the rate constants of 34 individual reactions taking place. Studies have also been conducted with peptides and proteins in which electrochemically-driven charge transfers have been demonstrated between methionine, tryptophan, tyrosine and cysteine/cystine units. Free radical reactions in Mitomycin C have been elucidated which are consistent with pulse radiolysis observations and the formation of radiolytic products as determined by HPLC. Adriamycin reduction has also been studied: the Adriamycin semiquinone is unusually stable with respect to dismutation but its lifetime is limited by a decomposition process in which daunosamine is expelled. The expulsion is followed by a further rearrangement. Many of the reactions investigated require tens or hundreds of seconds to reach essential completion.

Over a wide range of pH, the semiquinone free radicals formed by reduction of adriamycin exist as a form which is strongly stabilised by internal hydrogen bonding and resonance. They protonate with pKa = 2.9. Below this pH they exhibit absorption maxima at 430 nm (Emax = 13,200 dm3 mol-1 cm-1) and approximately 720 nm (Emax = 4,200 dm3 mol-1 cm-1). Above pH 2.9 they have maxima at 480 nm (Emax = 14,600 dm3 mol-1 cm-1) and approximately 700 nm (Emax = 3,400 dm3 mol-1 cm-1). In acid and alkaline solution the radicals rapidly disappear by disproportionation, but within the approximate pH range 6 to 11 they appear to be relatively stable for at least 10-20 ms, existing in transient equilibrium with parent adriamycin and the full reduced form. Some rate constants for the formation and reactions of the semiquinone are given, including the reaction with oxygen to give O2.-. Fully reduced adriamycin has absorption maxima at 410 nm (Emax = 11,000 dm3 mol-1 cm-1) at pH 5 and 430 nm (Emax = 19,000 dm3 mol-1 cm-1) at pH 11. It undergoes decomposition within a few hundred ms. The intermediates from daunomycin would be expected to have properties similar to those from adriamycin.

The OH radical rapidly reacts with CH3NH2 and C6H5NH2 but reacts slower with CH3NH3 ⊕, respectively C6H5NH3 ⊕. In the reaction with methylamine. the complex is formed which loses water with τ½=26 µs. The addition of OH to the aromatic ring of aniline occurs in 20% of the reactions. The hydroxy-amino-cyclohexadienyl radical has its absorption maximum at 3500 A. It decays by elimination of water to yield the anilino radical C6H5NH (τ½= 5.7µs at pH = 7), H⊕ and OH⊖ ions catalyse the decay. In 80% of the reactions. · OH adds to the amino group of aniline. The subsequent loss of water or of OH⊖ occurs rapidly ( is estimated to be equal to 11 — 12. The absorption spectra of C6H5NH2® and the pK-value of 7.0 of C6H5NH2 ⊕ first described by LAND and PORTER are confirmed.

WasBrige Losungen von Rhodamin B, Trypaflavin und alkalisch-wasrige Lo-sungen von Fluorescein (pH 13) zeigen nach Bestrahlung mit 2 Μsec Elektronenimpulsen von 4 MeV ein Nachleuchten, das bis zu 100 Μsec nach dem Impuls noch beobachtet werden kann. Die Lichtintensitat steigt nach dem Abklingen der Cerenkov-Strahlung zunachst bis zu einem Maximum an. Dieses ist bei Rhodamin B besonders ausgepragt und tritt bei den anderen Farbstoffen erst nach Zugabe von geringen Mengen KJ in Erscheinung. Das Spektrum des Nachleuchtens ist identisch mit dem durch UV angeregten Fluoreszenzspektrum. Bei [F] ≈ 2· 10−6M zeigt die Gesamtemission nach Messungen mit Rhodamin B ein Maximum. Die Quantenausbeute ist bei [F]=10−6M von der Grosenordnung 10−5 Quanten/eV. Durch wirksame e aq − -Fanger wird das Nachleuchten wirksam geloscht. Durch Zugabe von geringen Mengen KJ wird die Intensitat um ein Vielfaches gesteigert. Nach dem gegenwartigen Stand der Untersuchungen ist wahrscheinlich, das die Anregung durch die Reaktionen OH+F→Fox e aq − +Fox→F*+OH− und die Sensibilisierung durch J− durch die Reaktionen e aq − +F→Fred J−+OH→J+OH− J+Fred→F*+J− geschieht.

A new method for determining these coefficients has been developed. The method employs pulse radiolysis for triplet excitation and, by means of triplet-triplet ( T–T ) energy transfer, leads to a comparison between the extinction coefficients of the benzophenone ketyl radical (ф 2 ĊOH) and the triplet states of several aromatic compounds whose triplet levels are lower than that of benzophenone. Since the ketyl extinction coefficient can be determined independently, the actual T–T extinction coefficients can be obtained. Values of the extinction coefficient at the absorption maximum were obtained for anthracene (57200 m -1 cm -1 ), naphthalene (22600 m -1 cm -1 ), phenanthrene (21000 m -1 cm -1 ) and 1,2-benzanthracene (25100 m -1 cm -1 ), all relative to an extinction coefficient for the benzophenone ketyl maxi­mum around 540 nm determined to be 3220 m -1 cm -1 . For several other compounds the above method for various reasons could not be applied. In such cases estimates were obtained by energy transfer experiments, leading to comparisons between their triplet absorptions and those of the compounds already determined. Estimates were thus obtained for the maximum T–T extinction coefficients of benzophenone (10300 m -1 cm -1 ), biacetyl (6400 m -1 cm -1 ), diphenyl (35400 m -1 cm -1 ), and acridine (28800 m -1 cm -1 ). During the course of these experiments estimates were also obtained for the rates of the following reactions : e - aq. + benzophenone ( k ii ═ 3·0 × 10 10 m -1 s -1 ), OH˙ + benzophenone ( k ii ═ 8·7 × 10 9 m -1 s -1 ) and hydrogen abstraction by triplet benzophenone from cyclo­hexane, to give ф 2 ĊOH ( k ii ~ 8 × 10 5 m -1 s -1 ).

Pulse radiolysis of solutions of iodine in cyclo hexane resulted in the formation of a transient absorption with wavelength maximum at 3300 Å. This transient spectrum was assigned to the C 6 H 12 ... I' charge-transfer complex. The spectrum and extinction coefficients of the complex over the wavelength range 2500 to 6500 Å were obtained, the coefficient at 3300 Å being 2025 1. mole -1 cm -1 . Hydrogen iodide in cyclo hexane gave rise to the same transient absorption. Pulse radiolysis of cyclo hexane and cyclo hexyl iodide alone gave rise to different transient spectra attributed to the cyclo hexyl radical, absorbing in the ultraviolet, extinction coefficient 3401. mole -1 cm -1 at 2550 Å and the C 6 H 11 .. .I* charge-transfer complex, absorption maximum 3900 A, extinction coefficient ~ 10 3 to 10 4 1. mole -1 cm -1 , respectively. During the course of the investigations rate constants were obtained for the following reactions: C 6 H 11 + I 2 C 6 H 11 I + I', k II = 7 x 10 9 1. mole -1 s -1 , 2C 6 H 12 ...I' -> 2C 6 H 12 + I 2 , k II = 1.06 x 10 10 1. mole -1 s -1 , or 21' -> I 2 , a C 6 H 10 + C 6 H 12 2C 6 H 11 k 11a + k 11b =2.5 x 10 9 1. mole -1 s -1 bC 12 H 22 Dilute solutions of iodine (10 -4 M) in cyclo hexane yielded experimental evidence for neutral radical intermediates only. There were, however, indications that ionic radical intermediates occur in more concentrated solutions ( > 10 -3 M).

— –Pulse radiolysis has been used to excite the triplet states of β-carotene (τ# 9μ sec) and lycopene (τ= 8μsec) in hexane solution, both in the presence and absence of naphthalene as a triplet sensitiser. The absorption spectra of both triplets have been measured in the range 430–550 nm and have thus been extended into the region of the corresponding singlet absorptions. The overlap of the triplet and singlet spectra is discussed in relation to in vivo studies. Extinction coefficients of 1.3±0.1 × 105 l/mole cm for β-carotene triplet 515 nm and 3.9±0.2 × 105 l/mole cm for lycopene triplet at 525 nm were obtained. Isomerisation of the all-trans polyenes used was detected and preliminary measurements indicate that the yield of isomerisation was greater than the triplet yield. The rate of triplet energy transfer from naphthalene to β-carotene was estimated to be 1.5 × 1010 l/mole sec. The corresponding value for lycopene was 1.4× 1010 l/mole sec. The measured efficient quenching of triplet β-carotene by oxygen may occur by an energy transfer mechanism, leading to the formation of singlet oxygen (1Δg. This would suggest that the triplet energy level of β-carotene lies between 121 and 94 kJ mole-1.

— A radical transformation of the type Trṗ-TyrOH TrpH-TyrO, previously observed by pulse radiolysis, has been detected following 265 nm laser excitation of oxygen saturated neutral aqueous solutions of TrpH-TyrOH, TyrOH-TrpH and cyclo-TrpH-TyrOH. A similar process was observed after photoexcitation of the proteins α-lactalbumin and α-chymotrypsin.

Flash photolysis and pulse radiolysis techniques1 have contributed substantially to our understanding of the harmful and beneficial effects of radiation. The development of extremely short pulsed excitation sources means that time resolution is no longer a serious limitation to the application of these methods2,3. However, these methods have previously been restricted to transparent samples because changes in transmission of analysing light were measured following pulsed excitation. Recently4–6, we have established that photochemical processes within opaque and highly scattering systems can be studied by observing changes in diffusely-reflected analysing light following laser excitation. Many of the most important radiation-induced processes involve non-transparent, heterogeneous systems, for example, most biological systems. We report here the first successful ‘diffuse-reflectance pulse radiolysis’ experiments in which we demonstrate that time-dependent absorption spectra of short-lived species can now be measured for opaque samples following pulse radiolysis.

In order to study the relationship between certain photophysical and photochemical properties of furocoumarins and their photobiological activities, the quantum yields of the formation of excited triplet states, the capacity to generate singlet oxygen in vitro and the effect of oxygen on photoinduced cell killing and on the induction of cytoplasmic "petite" mutations in Saccharomyces cerevisiae were determined for a series of 4'-methylangelicins and 3-carbethoxypsoralen (3-CPs). The capacity to generate singlet oxygen, in good agreement with the photophysical data, correlates well with the oxygen effect observed on the lethality of Saccharomyces cerevisiae shown by the various compounds and followed the ranking order 3-CPs greater than 4',4-dimethylangelicin greater than 4'-methylangelicin greater than 4',5-dimethylangelicin. On the other hand the oxygen effect on the induction of cytoplasmic "petite" mutations on yeast appears much less pronounced than that shown on survival. Concerning other photobiological properties previously studied no correlation was observed between the capacity to generate singlet oxygen in vitro and the skin photosensitizing activity of the compounds. Moreover, the generation of singlet oxygen by furocoumarins does not appear as the main cause for the photoinduction of skin tumors in mice.

The semireduced, semioxidized, and OH(.)-adduct radicals of bilirubin (BR) and biliverdin (BV) have been characterized using pulse radiolysis techniques. Laser flash photolysis (265-nm) of these pigments led to monophotonic photoionization with quantum yields of 0.08 for BR and 0.03 for BV. No evidence for triplet formation or for photoisomerization was found after 265-nm laser excitation. However, 347-nm excitation of BR in chloroform led to simultaneous photoisomerization and radical formation, but the radicals are thought to have originated from a pathway other than photoionization. The relevance of these observations to BR photoreactivity is discussed. BR radical ions in alkaline solution did not react with tryptophan (TrpH), but the semioxidized TrpH radical oxidized BR with k = 4.3 X 10(8) dm3 mole-1 sec-1. When human serum albumin (HSA) was oxidized using radiolytically generated azide radicals, a radical transformation involving TrpH and TyrOH residues occurred with k = 3.8 X 10(3) sec-1. When BR was complexed with the protein the transformation rate was reduced to 1.6 X 10(3) sec-1. This was interpreted in terms of a conformational change in the protein. Identification of the probable residues involved provided information about the primary BR binding site which was consistent with an earlier report.

A series of radical transitions, Br2-.----Met(S therefore Br)----Trp(indolyl)----Tyr (phenoxyl), has been demonstrated by pulse radiolysis of N2O-saturated aqueous solutions containing Br-, Met-Gly and Trp-(Gly)2-Tyr at pH 6.7. The intramolecular Met(S therefore Br)----Trp(indolyl) transition in the dipeptide Met-Trp is shown to proceed via the Trp+. radical cation, with a rate constant of k approximately 10(7)s-1, consistent with an electron transfer. Br2-.-attack upon ribonuclease A (RNase) leads to a fast Met(S therefore Br)----Tyr(phenoxyl) process, k = (4.0 +/- 1.0) X 10(5)s-1, probably involving the solvent-exposed Met-29 and the adjacent Tyr-25. Phenoxyl dimerization in the RNase system produces the characteristic o,o'-biphenol fluorescence, but a competing interaction of the Tyr-25(phenoxyl) with the 26-84 disulphide group also appears possible.

In this paper the authors report the first successful time-resolved detection of singlet oxygen, O/sub 2/(/sup 1//Delta//sub g/), luminescence using the pulse radiolysis technique. The use of this technique (a) to produce high concentrations of solute (S) triplet states in aerated benzene (B) via a combination of channels 1-4 and (b) to subsequently form O/sub 2/(/sup 1//Delta//sub g/) via channel 5 has already been described. The method complements direct pulsed laser excitation of S in that formation of /sup 3/S*, and therefore of O/sub 2/(/sup 1//Delta//sub g/), is still efficient in those instances where intersystem crossing (channel 4) is unimportant. In the latter situation a laser-based experiment would require an additional light-absorbing sensitizer which could subsequently transfer triplet energy to high concentrations of S. Such experiments, certainly of a quantitative nature, are usually doomed to failure because of competitive light absorption problems. No such problems exist with pulse radiolysis, and the high available triplet energy of /sup 3/B* (84 kcal mol/sup /minus/1/) ensures that virtually any solute of interest, in the O/sub 2/(/sup 1//Delta//sub g/) context, will be efficiently promoted to its triplet state.

The active component of ‘haematoporphyrin derivative’ (HpD) is probably some aggregate involving both haematoporphyrin (Hp) and hydroxyethylvinyldeuteroporphyrin (HVD) at least as far as selective accumulation in tumours is concerned. Subsequent monomerisation in the cell could occur with the aggregate acting as a ‘pool’ of Hp and HVD so that it is of interest to examine the photophysical properties of such molecules in environments where they may be expected to be monomers as well as in environments where aggregation occurs. In this paper we describe the photophysical properties of Hp in acetone, various detergents, methanol-water mixtures, phosphate buffers and aqueous alkali as well as preliminary results using phosphatidyl choline (PC) vesicles and white blood cells. The major part of the work concerns the study of the lowest excited triplet state of Hp by laser flash photolysis (347 nm excitation).

A water soluble derivative of 4,5'-dimethylangelicin (I) having a long chain linking an amino group to the planar furocoumarinic moiety, that is 4'-N,N-dimethylaminoethoxymethyl-4,5'-dimethylangelicin (III), has been prepared. This compound is able to form effectively the intercalated complex with DNA like the previously prepared 4'-aminomethyl-4,5'-dimethylangelicin (II), however while the compound (II) showed very poor photobinding to DNA, the new derivative (III) shows high photobinding to the macromolecule. It is proposed that these data illustrate the importance of the geometry of intercalation for the subsequent covalent photobinding to the macromolecule. Also some photophysical data of (II) and of (III) appear to confirm the critical role of the position assumed by the chromophore of the two compounds when intercalated in duplex DNA. The compound (III) displays high photobiological effects, also in terms of antiproliferative activity as shown by its capacity to inhibit DNA and RNA synthesis in Ehrlich cells, the growth of an E. coli culture and the infectivity of T2 phage. (III) on the basis of these properties seems to deserve a clinical evaluation of its potential photochemotherapeutic activity in the treatment of psoriasis.